标准液中颜色检测方案(浊度计)

上海胤煌科技让检测分析更专业 中/美/欧/日四大药典澄清度检查规范-中英双译 中国药典20200902澄清度检查法 澄清度检查法系将药品溶液与规定的浊度标准液相比较，用以检查溶液的澄清度。除另有规定外，应采用第一法进行检测。 品种项下规定的“澄清”，系指供试品溶液的澄清度与所用溶剂相同，或不超过0.5号浊度标准。“几乎澄清”，系指供试品溶液的浊度介于0.5号至1号浊度标准液的浊度之间。 第一法(目视法) 除另有规定外，按各品种项下规定的浓度要求，在室温条件下将用水稀释至一定浓度的供试品溶液与等量的浊度标准液分别置于配对的比浊用玻璃管(内径15-16mm，平底，具塞，以无色、透明、中性硬质玻璃制成)中，在浊度标准液制 备5分钟后，在暗室内垂直置于伞棚灯下，照度为1000lx， 从水平方向观察、比较。除另有规定外外，供试品溶解后应立即检视。 第一法无法准确判定两者的澄清度差异时，改用第二法进行测定，并以其测定结果进行判定。 浊度标准存贮液的制备称取于105℃干燥至恒重的硫酸肼1.00g，置于100ml量瓶中，加水适量使溶解，必要时可在40℃的水浴中温热溶解，并用水稀释至刻度，摇匀，放置4-6小时；取此溶液于等容量的10%乌洛托品溶液混合，摇匀，于25℃避光静置24小时，即得。该溶液置冷处避光保存，可在2个月内使用，用前摇匀。 浊度标准原液的制备取备度标准贮备液15.0ml， 置1000ml量瓶中，加水稀释至刻度，摇匀，取适量，置1 cm吸收池中，照紫外-可见分光光度法(通则0401)， 在 550 nm 的波长处测定，其吸光度在0.12-0.15范围内，该溶液应在48小时内使用，用前摇匀。 浊度标准液制备取浊度标准原液与水，按照下表配置，即得。浊度标准液应临用时制备，使用前充分摇匀。 级号 0.5 1 2 3 4 浊度标准原液(ml) 水 (ml) 2.5097.50 5.095.0 10.090.0 30.070.0 50.050.0 第二法(浊度仪法) 供试品的浊度可采用浊度仪测定。溶液中不同大小、不同特性的微粒物质包括有色物质均可使入射光产生散射，通过测定透射光或者散射光的强度，可以检 ( Email/邮箱： info@yh-t e k.co m P hone/电话：021-58220558 V Website/网址： w ww.y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) 查供试品的浊度。仪器测定模式通常有三种类型，透射光式、散射光式和透射光-散射光比较测量模式(比率浊度模式)。 1.仪器的一般要求 采用散射光式浊度仪时，光源峰值波长为860 nm；测量范围应包含 0.01-100NTU。在 0-10NTU 范围内分辨率应为 0.01NTU； 在 10-100NTU 范围内分辨率应为 0.1NTU. 2.适用范围及检测原理 本法采用散射光式浊度仪，适用于低、中浊度无色供试品溶液的浊度测定(浊度值为100NTU 以下的供试品。)因为高浊度的供试品会造成多次散射现象，时 散射光强度迅速下降，导致散射光强度不能正确反映供试品的浊度值。0.5-4号浊度标准液的浊度值范围约为 0-40NTU. 采用散射光式浊度仪测定时，入射光和测定的散射光呈90℃夹角，入射光强度和散射光强度关系式如下。 式中Ⅰ为散射光强度，单位为 cd； Io 为入射光强度，单位为 cd； K'为散射系数： T为供试品溶液的浊度值，单位为NTU (NTU 是基于福尔马肼浊度标准液液测定的散射浊度单位，福尔马肼浊度标准液即为第一法中的浊度标准贮备液)。 在入射光Io不变的情况下，散射光强度Ⅰ与浊度值成正比。因此，可以将浊度测量转化为散射光强度的测量。 3.系统的适用性试验 仪器应定期(一般每月一次)对浊度标准液的线性和重复性进行考察，采用0.5号至4号浊度标准液进行浊度值测定，浊度标准液的测定解果(单位NTU)与浓度间应呈线性关系，线性方程的相关系数应不低于0.999；取0.5号至4号浊度标准液，重复测定5次，0.5号和1号浊度标准液测量浊度值的相对标准偏差应不大于5%，2-4号浊度标准液测量浊度值的相对标准偏差不大于2%。 4.测定法 按照仪器说明书要求并采用规定的浊度液进行仪器校正。溶液剂直接取样测定；原料药或者其它剂型按照个论项下的标准规定制备供试品溶液，临用时制备。分别取供试品溶液和相应浊度标准液进行测定，测定前应摇匀，并避免产生气泡，读取浊度值。供试品溶液浊度值不得大于相应浊度标准液的浊度值。 美国药典USP44 <630> VISUAL COMPARISON 视觉比较 The purpose of this test is to provide the details for the visual comparison of thecolor and/or turbidance of sample solutions of certain concentration to a standardsolution or a series of standard solutions of known concentration. Where a color or turbidity comparison is directed， follow the procedures and conditions outlined belowfor performing these tests. 本试验的目的是提供特定浓度的样品溶液与已知浓度的标准溶液或一系列标准溶液的颜色和/或浊度的视觉比较细节。如果需要进行颜色或浊度比较，请遵循以下程序和条件进行这些测试 Comparison vessels： Color-comparison tubes matched as closely as possible ininternal diameter， in depth of sample solution， and in all other respects should be used.对比容器：应使用内径、样品溶液深度和所有其他方面尽可能匹配的颜色对比管。 Viewing conditions for turbidity comparison： Tubes should be viewedhorizontally against a dark background with the aid of a light source directed from thesides of the tubes. 浊度比较的观察条件：应在黑暗背景下，借助从管子侧面发出的光源水平观察管子。 Viewing conditions for color comparison： Tubes should be viewed downwardagainst a white background. Most of the time， common room lighting is sufficient toperform the assessment. A light source directed from beneath the bottoms of the tubesmay be used if needed and if the practice is consistent between the materials undercomparison. 颜色比较的观察条件：管子应在白色背景下向下观察。大多数情况下，公共空间照明足以进行评估。如果需要，并且对比材料之间的实践一致，可以使用从管底部下方引导的光源 <855> NEPHELOMETRY AND TURBIDIMETRY散射光浊度法和透射光比浊法 1.INTRODUCTION 介绍 Nephelometry and turbidimetry are analytical techniques that are based on theprinciples of light-scattering phenomena. Light scattering is the physical phenomenonin which a beam of light changes its direction of propagation (known as deflection) asa result of interaction with sufficiently small matter particles. It has been establishedfrom the Maxwell electromagnetic theory that a prerequisite for scattering to occur isthat the refractive indexes of the suspended particles must be different from those ofthe suspending liquid. The larger the difference， the more intense the scatteringbecomes. There are two types of light scattering： 1) elastic scattering， in which thewavelength of the scattered light and incident light are the same； and 2) inelastic light ( Email/邮箱： info@ y h-tek.c o m Phone/电话： 0 21-58220558 W V ebsite/网址： w ww.y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) scattering， in which the wavelength of the scattered light and incident light are different.Only the first type of light scattering (elastic) is relevant to nephelometry andturbidimetry. 散射光浊度法和透射光比浊法是基于光散射现象原理的分析技术。光散射是一种物理现象，其中光束由于与足够小的物质粒子相互作用而改变其传播方向(称为偏转)。根据麦克斯韦电磁理论，散射发生的先决条件是悬浮颗粒的折射率必须不同于悬浮液体的折射率。差异越大，散射越强烈。光散射有两种类型：1) 弹性散射，其中散射光和入射光的波长相同；2)非弹性光散射，其中散射光和入射光的波长不同。只有前一种光散射(弹性)与散射光浊度法和透射光比浊法有关。 In turbidimetry， the intensity of the transmitted light is measured and theattenuation of the intensity of incident light as a result of scattering is measured at thedirection of incident light (i.e.， 0°) and compared to the intensity of incident light (blankmeasurement). The measured property is an indirect measurement of the scatteringeffect of the suspended particles and is referred to as turbidance. Any absorbance oflight by the suspended sample will result in additional attenuation of light intensity (seeUltraviolet-Visible Spectroscopy <857> andUltraviolet-Visible Spectroscopy-Theory and Practice <1857>). Hence， it is important to ensure that the material beingmeasured does not absorb light at the measurement wavelength. Indeed the equationsgoverning absorption and turbidimetry are the same (albeit with different values for theattenuation constants). In nephelometric techniques， the intensity of the scattered lightat a 90° angle from the propagation direction of the incident light is measured.Therefore， a nephelometric measurement is a direct measurement of the scatteringeffect of suspended matter. 在透射光比浊法中，测量透射光的强度，并在入射光方向(即 0°)测量散射导致的入射光强度的衰减，并与入射光强度进行比较(空白测量)。被测特性是悬浮颗粒散射效应的间接测量，称为浊度。悬浮样品对光的任何吸收都会导致光强度的额外衰减(参见<857>Ultraviolet-Visible Spectroscopy 和<1857> Ultraviolet-Visible Spectroscopy-Theory and Practice)。因此，确保被测材料不会吸收测量波长处的光非常重要。实际上，控制吸收和浊度测定的方程式是相同的 (尽管衰减常数的值不同)。在散射光浊度法中，测量与入射光传播方向成90°角的散射光强度。因此，散射光浊度法浊度测量是对悬浮物散射效应的直接测量。 2.TERMS AND DEFINITIONS 术语和定义 Terms commonly used in describing turbidimetric and nephelometric techniques are： · Turbidance (symbol， S)： A measure of the decrease of the transmitted incident lightbeam intensity as a result of the light-scattering effect of suspended particles. Theamount of suspended matter may be measured by observation of either thetransmitted light (turbidimetry) or the scattered light (nephelometry). Io= intensity of incident light It=intensity of transmitted light 上海胤煌科技让检测分析更专业 k= molar turbidity coefficient b = cell path length c=concentration T=turbidance ·Turbidity (symbol， t)： In turbidimetric measurements， the turbidity is the measure ofthe decrease in incident beam intensity/unit length ofa given suspension. TheInternational Organization for Standardization defines turbidity as “the reduction oftransparency of a liquid caused by the presence of undissolved matter”. · Turbidity Measurement Units： The turbidity units are stated using a descriptor whichindicates the method of measurement. ·Nephelometric Turbidity Units (NTUs)： When the turbidity is measured using anephelometer， which measures the scattered light at a 90° angle from the directionof propagation of incident light， the units of turbidity are called nephelometricturbidity units (NTUs). The magnitude of NTU is defined based on the turbiditygenerated by primary formazin standard (a suspension made by mixing solutions ofhydrazine sulfate and hexamethylenetetramine in water). Safer polymer-beadsuspensions are now commercially available and are recognized as an acceptablealternative. However， all those standards are traced to formazin. The primaryformazin standard solution has been assigned a turbidity of 4000 NTUs. Other recognized units for turbidity include the formazin turbidity unit (FTU) andthe formazin nephelometric unit (FNU). These units are equivalent to NTU for therange from 0-40 NTUs. 描述浊度法和浊度法的常用术语包括： 浊度(符号S)：由于悬浮颗粒的光散射效应，透射入射光束强度降低的一种度量。悬浮物的量可以通过观察透射光(比浊法)或散射光(浊度法)来测量O lo=入射光强度 It=透射光强度 k=摩尔浊度系数 b=样品池路径长度 c=浓度 T=浊度 ·浊度(符号，Tt)：在透射光浊度测量中，浊度是给定悬浮液的入射光束强度/单位长度减少的量度。国际标准化组织将浊度定义为“由于存在未溶解物质而导致液体透明度降低”。 .浊度测量单位：浑浊度单位用一个描述符表示，该描述符指示测量方法。 ·散射光浊度计浊度单位(NTU)：当使用散射光浊度法测量浊度时，浊度计以与入射光传播方向成90°角的角度测量散射光，浊度单位称为散射光浊度法浊度单位(NTU)。NTU 的大小是根据初级福尔马标准品(一种将硫酸肼和六亚甲基四胺溶液混合在水中制成的悬浮液)产生的浊度定义的。更安全的聚合物微珠悬浮液现已上市，并被公认为可接受的替代品。然而，所有这些标准都可以追溯到福尔马肼。初级福尔马肼标准溶液的浊度为4000NTU。 其他公认的浊度单位包括福尔马肼比浊法单位(FTU)和福尔马肼浊度法单位(FNU)。这些单位相当于0-40 NTU 范围内的 NTU。 3.APPLICATIONS 应用 Turbidimetric and nephelometric techniques have applications that include 1)concentration determination of solutions and/or suspensions (determination of severalcations and anions by precipitating and suspending the resulting precipitate at well-controlled reaction parameters)； 2) measurement of the degree of turbidity of turbidsolutions or suspensions； 3) determination of weight-average molecular weights anddimensions of polydisperse systems in the molecular weight range from 1000 to severalhundred million； 4) measurement of immunoassays’reaction kinetics or kinetics ofimmunoprecipitations (rate nephelometry)；5) monitoring of cell and bacteria growth；and 6) particle size distribution determination of suspended material， particle counting，etc. 透射光比浊法和散射光浊度法技术的应用包括1)溶液和/或悬浮液的浓度测定(通过在控制良好的反应参数下沉淀和悬浮产生的沉淀物，来测定几种阳离子和阴离子)；2)测量混浊溶液或悬浮液的浊度；3)测定分子量在1000到数亿之间的多分散体系的重均分子量和尺寸；4)测量免疫分析的反应动力学或免疫沉淀动力学(比率散射浊度法)；5)监测细胞和细菌的生长；6)悬浮物粒度分布测定、颗粒计数等。 Rate nephelometry is widely used for vaccine componentss assays and/orquantitation of components in blood serum. It is also used for host cell proteinqualification in recombinant biopharmaceuticals. When using the technique， themeasurement of the change in the light-scattering response by antigen-antiserum orantigen-purified antibody complexes is used to calculate the amount of antigen (Ag) orantibody (Ab) responsible for the immunological Ab-Ag precipitation reaction oragglutination reaction. Often the antigens under consideration are linked covalently oradsorbed to polymeric microspheres to increase the scattering efficiency； the resultingtechnique is known as "particle-enhanced immunoassay". Although the technique isdescribed as nephelometry， usually both scattered and transmitted light are measuredusing the ratio instruments. 比率散射浊度法广泛用于疫苗成分分析和/或血清成分的定量。它还用于重组生物制药中的宿主细胞蛋白质鉴定。当使用该技术时，通过测量抗原-抗血清或抗原纯化抗体复合物的光散射反应的变化，来计算导致免疫抗体-抗原沉淀反应或凝集反应的抗原(Ag) 或抗体(Ab)的量。通常考虑抗原共价连接或吸附在聚合物微球上，以提高散射效率；由此产生的技术被称为“颗粒增强免疫分析”。虽然这项技术被称为散射光浊度法，但通常散射光和透射光都是用比率仪器测量的。 Nephelometric measurements are more reliable in low turbidity ranges (relativelylow concentration of the scattering medium). In this range， a linear relationship isobserved between the sample concentration and the detector’s signal intensityexpressed as NTU. As the concentration increases， so does the incidence of multiplescattering that deviates the response from the linearity. The maximum NTU value，which supports a reliable linearity relationship， is in the range of 1750-2000 NTUs. ( Email/邮箱： info@ y h-tek.co m Phone/电话： 0 21-58220558 W V ebsite/网址： w ww.y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) Turbidimetry is preferred for higher turbidity ranges (concentrations ofthe scatteringmedia). To achieve consistent results， all measurement variables must be carefullycontrolled. Where such control is possible， extremely dilute suspensions may bemeasured. 散射光法浊度测量在低浊度范围(散射介质浓度相对较低)更可靠。在该范围内，观察到样品浓度与检测器信号强度(以NTU 表示)之间存在线性关系。 随着浓度的增加，多次散射的入身角也会增加，从而偏离线性响应。支持可靠线性关系的最大NTU 值在1750-2000 NTU 范围内。透射光比浊法适用于更高的浊度范围(散射介质的浓度)。为了获得一致的结果，必须仔细控制所有测量变量。在可能的情况下，可以测量极稀的悬浮液。 4.INSTRUMENTATION 仪器仪表 Instruments used for turbidimetric and nephelometric measurements are calledturbidimeters and nephelometers， respectively. Generally， these instruments consist ofa mercury lamp with filters for the strong green or blue lines， a shutter， a set of neutralfilters with known transmittance， and a sensitive photomultiplier， which can bemounted fixed at 0° or at a 90° angle from the incident light propagation direction， oron an arm that can be rotated around the solution cell and set at any angle from-135°to 0° to +135° by a dial outside of the light-tight housing. Solution cells are of variousshapes， such as square for measuring 90° scattering； semioctagonal for 45°， 90°， and135°scattering； and cylindrical for scattering at all angles (see Figure 1). 用于透射光比浊法和散射光浊度法测量的仪器分别称为透射光浊度计和散射光浊度计。通常，这些仪器包括一个带有滤光器的汞灯(用于强绿线或蓝线)、一个快门、一组具有已知透射率的中性滤光器和一个灵敏的光电倍增管，该光电倍增管可安装在与入射光传播方向成0°或90°角的位置，或者在一个臂上，它可以围绕溶液单元旋转，并通过不透光外壳外的表盘设置为-135°到0°到+135°的任何角度。溶液池的形状多种多样，例如用于测量90°散射的正方形；45°、90°和135°散射为半八角形；圆柱形可适用于所有角度的散射(见图1)。 Detector 2 Figure1. Representative nephelometric (turbidimetric) instrument. Note that Detector2 may be mounted on a movable arm. 图1。代表性浊度仪。注意，探测器2可安装在可移动臂上。 Turbidity also can be measured with a standard photoelectric filter photometer orspectrophotometer， preferably with illumination in the blue portion of the spectrum.Nephelometric measurements require an instrument with a photocell placed so as toreceive scattered， rather than transmitted， light. Because this is the same geometry usedin fluorometers， they can be used as nephelometers by proper selection of filters. A ratioturbidimeter combines the technology of 90° nephelometry and turbidimetry. Itcontains photocells that receive and measure scattered light at a 90° angle from thesample as well as receiving and measuring the forward scatter in front of the sample. Italso measures light transmitted directly through the sample. Linearity is attained bycalculating the ratio of the 90° angle scattered light measurement to the sum of theforward scattered light measurement and the transmitted light measurement. Thebenefit of using a ratio turbidimetric system is that the measurement of stray lightbecomes negligible. In addition， the determination of turbidity of colored suspensionsis done exclusively using turbidimetric or nephelometric instruments with ratio modebecause this procedure compensates for the attenuation of light as the result of thesuspension color. Typically， the light source in these instruments is a tungsten lampwith most of the light intensity at about 550 nm operating at the filament temperatureof 2700 K. Other suitable light sources are also available. Typically， the detectors are▲silicon diodes(ERR 1-May-2019) and photomultipliers. An alternative for eliminatingthe color effect involves using an infrared light-emitting diode as a light source， whichyields an emission maximum centered at about 860 nm and a spectral bandwidth of 60nm. When laser light sources are also used， especially in nephelometric instruments，the technique is commonly known as "laser nephelometry". The advantage of usinglaser nephelometers is the significant improvement in signal-to-noise ratio at very lowdetection levels. Usually the light source is a laser diode with a working wavelength at660 nm. The high-power density of the laser beam gives rise to higher scatteredintensity from smaller particles. Combined with a light trap， which absorbs theunscattered light， the system lowers the stray light significantly. When the use of anephelometer or turbidimeter is indicated for a procedure in a monograph， instrumentsworking in ratio mode may be used instead. 浊度也可以用标准光电滤光光度计或分光光度计测量，最好是在光谱的蓝色部分进行照明。散射光浊度法测量需要一个装有光电管的仪器，以便接收散射光，而不是透射光。由于这与荧光计中使用的几何结构相同，因此可通过适当选择滤光片将其用作浊度计。比率浊度计结合了90°散射光浊度法和透射光比浊法。它包含光电管，接收和测量与样品成90°角的散射光，以及接收和测量样品前面的前向散射光。它还测量直接穿过样品的光。通过90°角散射光测量值，前向散射光测量值和透射光测量值之和，计算两者的比值，可获得线性度。使用比率浊度测量系统的好处是杂散光的测量变得可以忽略不计。此外，彩色悬浮液的浊度测定仅使用透射光比浊法浊度仪或浊度仪(带比率模式)进行，因为该程序补偿了悬浮液颜色导致的光衰减。通常，这些仪器中的光源是钨灯，在2700K的灯丝温度下工作，大部分光强约为550nm。也可使用其他合适的光源。通常，探测器是▲硅二极管▲和光电倍增管。另一种消除颜色效应的方法是使用红外发光二极管作为光源，其最大发射中心约为860nm，光谱带宽为60nm。当激光光源也被使用时，尤其是在浊度测量仪器中，这种技术通常被称为“激光浊度测量”。使用 ( Email/邮箱： info@ y h-tek.c o m Phone/电话： 0 21-58220558 W V ebsite/网址： w ww.y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) 激光散射光浊度计的优点是，在非常低的检测水平下，信噪比显著提高。通常光源是工作波长为660 nm 的激光二极管。激光束的高功率密度使交小粒子产生更高的散射强度。与吸收未散射光的光阱相结合，该系统可显著降低杂散光。当专著中的某个程序指示使用散射光浊度计或透射光浊度计时，可以使用在比率模式下工作的仪器。 5.FORMAZIN TURBIDITY STANDARDS 福尔马肼浊度标准 Formazin is the only known primary turbidity standard. All other standards aresecondary and must be traced to formazin. The primary standard is defined in the▲IUPAC Compendium of Chemical Terminology，▲ (ERR 1-May-2019) 2nd ed. (theGold Book) as one that is prepared by the user from traceable materials using well-defined methodologies and conditions. 福尔马肼是唯一已知的主要浊度标准。 所有其他标准都是次要的，必须追溯到福尔马肼。主要标准在▲IUPAC Compendium of Chemical Terminology▲(ERR 1-May-2019) 第2版(金书)中被定义为由用户使用明确定义的方法和条件从可追溯的材料准备的标准。 Formazin suspension has many features that ensure its suitability as a primarystandard. It can be consistently and accurately prepared from reagent-grade startingmaterials. The suspension consists of random polymers with different lengths and ofrandom configurations， which result in moieties of varying shapes and sizes rangingfrom less than 0.1 um to more than 10 um. Although the polymer chain lengthdistribution has been shown to vary from preparation to preparation， the overallresulting turbidity has been very reproducible. 福尔马肼悬浮液有许多特点，以确保其适合作为主要标准。它可以从试剂级的起始材料中始终如一、准确地制备。该悬浮液由不同长度和随机构型的聚合物组成，其组成的聚合物的形状和尺寸从小于 0.1 um 到大于 10 um 不等。尽管聚合物链长分布已被证明因制备而异，但总的浊度结果是可以很好地重现的。 1. Preparation of the Formazin Standards 福尔马肼标准液的制备 [NOTE-All procedures described below must be performed at 20 ± 2°(seeVolumetric Apparatus <31>.] Hydrazine sulfate solution： Dissolve 1.000 g of ACS grade hydrazine sulfate(N2H4H2SO4) in particle-free water in a 100-mL Class A volumetric flask and dilutewith particle-free water to volume. Allow this solution to stand for 4-6h. · Primary formazinstandard：Dissolve22.50 goffanalyticalgradehexamethylenetetramine [(CH2)6N4] in 25.0 mL of particle-free water in a 100-mL flask.Add 25.0 mL of hydrazine sulfate solution using a Class A 25-mL“to deliver”pipetteand mix thoroughly. Allow the preparation to stand for 48 h at 25±1° before using.The resulting suspension is stable for 2 months. ·Formazin stock standard suspension 1： Using a 15-mL Class A“to deliver”pipette，transfer 15 mL of the Primary formazin standard to a 1-Lvolumetric flask， and dilute with particle-free water to volume and mix. The resulting suspension has a turbidity of60 NTU. ·Formazin stock standard suspension 2： Using a 50-mL Class A“to deliver” pipette，transfer 50 mL of Primary formazin standard to a 200-mLvolumetric flask， and dilutewith particle-free water to volume and mix. The resulting suspension has a turbidity of1000 NTUs. ·Formazin reference suspensions： Prepare by mixing in a 100-mL volumetric flask，portions of the respective Formazin stock standard suspension and particle-free wateraccording to Table 1. [注： L以下所有的程序必须在20±2°的条件下进行(参见<31VolumetricApparatus>) ] •硫酸肼溶液：将1.000 g ACS 级硫酸酒 (N2H4H2SO4)溶解在100mL的A 类容量瓶中中，并用无颗粒水稀释至刻度。让该溶液静置4-6小时。 ·初级福尔马标准液：将2.50g分析级六亚甲基四胺[(CH2)6N4]溶于 25.0 mL无颗粒水中，装入100 mL烧瓶。使用A级25ml移液管加入25.0 mL 硫酸肼溶液，并充分混合。使用前，让制剂在25±1°的温度下静置48小时。由此产生的悬浮液可稳定运行2个月。 福尔马肼储备标准悬浮液1：使用15mLA 类移液管，将15 mL福尔马肼初级标准液转移至1L容量瓶中，并用无颗粒水稀释至刻度并混合。所得悬浮液的浊度为60 NTU。 ·福尔马肼储备标准悬浮液2：：使用 50mLA类移液管，将50mL 福尔马肼初级标准液转移至200mL容量瓶中，并用无颗粒水稀释至刻度并混合。所得悬浮液的浊度为1000 NTU。 ·福尔马肼参考悬浮液：根据表1，在100mL容量瓶中混合各份福尔马肼储备标准悬浮液和无颗粒水，制备福尔马肼参考悬浮液。 Table 1. Formazin Reference Suspensions Preparations Formazin ReferenceSuspensions Formazin Stock StandardSuspension 1Volume(mL) Formazin Stock StandardSuspension 2Volume (mL) Particle-Free WaterVolume(mL) Turbidity Value(NTU) Reference suspension I 5.0 95.0 3 Reference suspension II 10.0 90.0 6 Reference suspension III 30.0 70.0 18 Reference suspension IV 50.0 一 50.0 30 Stock standard suspension1 100 0 60 Reference suspension VI 10.0 90.0 100 Reference suspension VII 20.0 80.0 200 Reference suspension VIII 40.0 60.0 400 Reference suspension IX 70.0 30.0 700 Stock standard suspension 2 一 100 0 1000 6. QUALIFICATION OF TURBIDIMETERS AND NEPHELOMETERS 透射光式浊度仪与散射光式浊度仪的鉴定 The suitability of a specific instrument for a given procedure is ensured by astepwise life cycle evaluation for the desired application from selection to instrumentretirement. The qualification comprises three components： 1) installation qualification ( Email/邮箱： info@ y h-tek.c o m Phone/电话： 0 21-58220558V Website/网址： w ww.y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) (IQ)， 2) operational qualification (OQ)， and 3) performance qualification (PQ) (seeAnalytical Instrument Qualification<1058>). 特定仪器对给定程序的适用性由从选择到仪器报废的预期应用的逐步生命周期评估来确保。鉴定包括三个部部：1)安装鉴定(IQ)、2)操作鉴定(OQ)和：3)性能鉴定(PQ)(参见<1058>Analytical InstrumentQualification 章节)。 The purpose of this section is to provide test methods and acceptance criteria toensure that the instrument is suitable for its intended use(OQ)， and that it will continueto function properly over extended time periods (PQ). As with any spectrometric device，a turbidimetric and nephelometric spectrometer must be qualified for both wavelength(x-axis， if not fixed) and photometric (y-axis， or signal axis) accuracy and precision，and meet the requirements for the stray light. OQ is carried out across the operationalranges required within the laboratory for both the absorbance and wavelength scales. 本节的目的是提供测试方法和验收标准，以确保仪器适合其预期用途(OQ)， 并在延长的时间段(PQ)内继续正常工作。与任何光谱仪一样，透射光式和散射光式浊度光谱仪必须具备波长(x轴，如果不固定)和光度(y轴或信号轴)的准确度及精度，并满足杂散光的要求。OQ是在实验室内吸光度和波长标度所需的操作范围内进行的。 Acceptance criteria for critical instrument parameters that establish “fitness forpurpose"are verified during IQ and OQ. Specifications for particular instruments andapplications can vary depending on the analytical procedure used and the desiredaccuracy of the final result. Instrument vendors often have samples and test parametersavailable as part of the IQ/OQ package. 在IQ和OQ期间， 验证确定“用途适用性”的关键仪器参数的验收标准。特定仪器和应用的规格可能因使用的分析程序和最终结果的预期准确度而异。仪器供应商通常将样品和测试参数作为 IQ/OQ包的一部分提供。 Wherever possible in the procedures detailed as follows， primary referencestandards or certified reference materials (CRMs) are to be used. Formazin is the onlyprimary reference standard used in turbidimetry and nephelometry. All the otherstandards， including the CRMs， must be correlated to formazin. The CRMs should beobtained from a recognized accredited source and include independently verifiedtraceable value assignments with associated calculated uncertainty. CRMs must be keptclean and free from dust. Recertification should be performed periodically to maintainthe validity of the certification. 在以下详述的程序中，应尽可能使用主要参考标准或认证参考材料(CRM)。福尔马肼是比浊法法和浊度法中唯一使用的主要参考标准。所有其他标准，包括 CRM， 必须与福尔马肼相关。CRM )应从认可的认证来源获得，并包括独立验证的可追溯值分配及相关的计算不确定性。CRM必须保持清洁，无灰尘。应定期进行重新认证，以保持认证的有效性。 6.1Calibration 校准 All of the turbidimetric and nephelometric instruments are calibrated againststandards of known turbidity. The instrument must be calibrated using formazinturbidity standards prior to its first time use and at least every 3 months or as specifiedby the vendor. Calibration is performed using at least four formazin turbidity standardswhose turbidity proportionally covers the range of interest. Many instrumentmanufactures provide calibration verification standards. They usually consist of sealedsample cells filled with a latex suspension or with metal oxide particles in polymer gel.These standards must be used only for checking the calibration in the time intervalsbetween the instrument recommended calibrations. 所有透射光式浊度仪和散射光式浊度仪均根据已知浊度的标准进行校准。在首次使用之前，必须使用福尔马肼浊度标准液对仪器进行校准，至少每3个月或按照供应商的规定进行一次校准。使用至少四种福尔马肼浊度标准液进行校准，其浊度按比例覆盖感兴趣的范围。许多仪器制造商提供校准验证标准。它们通常由其中充满聚合物凝胶中的金属氧化物颗粒的密封样品池或乳胶悬浮液组成。这些标准只能用于检查仪器推荐校准的时间间隔内的校准。 2.. Stray Light 杂散光 Stray light (stray radiant energy) is a very significant error source， especially formeasurements in the range of the lower turbidity readings. It is defined as external lightthat reaches the detector without being scattered from the sample. There are severalsources of stray light including the inherent cell surface imperfections， reflections fromwithin the cell that are unaccounted for， optical system parts，light sources， and， to asmaller degree， the electronics fluctuations. Although there are many design featuresthat instrument vendors use to minimize the stray light， a complete mitigation of thestray light cannot be achieved. Unlike spectrophotometric measurements， the stray lightcannot be compensated for in turbidimetry. The stray light must be measured and thevalues should be within the specification range set by the vendor of the particularinstrument or <0.15 NTUs for the measurement in the range of 0-10 NTUs and 0.5NTUs for the measurements in the range of 10- 1100 NTUs， whichever is smaller. 杂散光(杂散光辐射能)是一个非常重要的误差源，特别是在较低的浊度读数范围内的测量。它被定义为到达探测器而不被样品散射的外部光线。杂散光有几种来源，包括电池表面固有缺陷、电池内部未被解释的反射、光学系统部件、光源，以及在较小程度上的电子波动。尽管仪器供应商使用了许多设计功能来最小化杂散光，但无法完全缓解杂散光。与分光光度测量不同，浊度法无法补偿杂散光。必须测量杂散光，其值应在特定仪器供应商设定的规格范围内， 或在0-10NTU 范围内测量时小于 0.15 NTU，在10-1100NTU 范围内测量时小于0.5 NTU，以较小者为准。 3.Range of Measuring Capability 测量能力的范围 The instrument must be able to measure the turbidity in the range of 0.01-1100NTUs or from 50%-200% of the target turbidity. To demonstrate the linearity for theintended measurements range， choose at least four appropriate reference suspensionsfrom Table 1. 上海胤煌科技让检测分析更专业 仪器必须能够测量 0.01-1100 NTU 范围内或目标浊度 50%-200%范围内的浊度。为了证明预期测量范围的线性， 从表 1中选择至少四种合适的参考悬浮液 6.4 Resolution 解决方案 Instrument resolution must be 0.01 NTU or less for the measurements range of 0-9.99 NTUs； 0.1 NTU or less for the measurements range of 10-99.9 NTUs； and 1NTUfor the measurements above 100 NTUs. 对于 0-9.99 NTU 的测量范围，仪器分辨率必须小于等于 0.01 NTU； 测量范围为 10-99.9 NTU 时，小于等于 0.1 NTU； 100 NTU 以上的测量分辨率值为1NTU. 5.. Accuracy准确度 The instrument reading accuracy must be ±10% of the reading+ 0.01 NTU for themeasurement range from 0-19.9 NTUs， and ±7.5% of the reading for the measurementrange from 20-1100 NTUs. 对于 0-19.9 NTU 的测量范围，仪器读数准确度必须为读数+0.01 NTU 的±10%， 对于 20-1100 NTU 的测量范围，仪器读数准确度必须为读数的±7.5%。 6. Performance Qualification 性能鉴定 The instrument PQ is accomplished periodically or as needed between thecalibrations. Primary turbidity standards(formazin) or secondary calibrationverification standards (latex suspensions or metal oxide particles in polymer gelscontained in sealed sample cells) supplied by instrument manufacturers may be used.1. 定期或根据需要在校准期之间完成仪器 PQ。可使用仪器制造商提供的一级浊度标准(福尔马肼)或二级校准验证标准(乳胶悬浮液或密封样品池中聚合物凝胶中的金属氧化物颗粒)。 7.PROCEDURE 步骤 1.. Turbidimetric Procedures 透射光比浊法测试步骤 SAMPLE CELL PREPARATION 样品池准备 The sample cells for sample measurements must be clean. Follow the sample cellor instrument manufacturer recommendationsfor cleaning the sample cellsappropriately. For low turbidity measurements it is a good practice to use a single-indexed sample cell or a flow cell， which help ensure adequate precision andrepeatability of the measurements. Using particle-free water， find the sample cellorientation in the sample cell holder that gives the lowest reading. For higher values ofturbidity， different sample cells may be used. However， the sample cells must bematched (the difference in readings for a standard prepared at nominal sampleconcentration from two different sample cells must be within ±0.005 NTU or below themeasurement precision requirement， whichever is lower). 用于样品测量的样品室必须清洁。按照样品池或仪器制造商的建议适当清洁样品池。对于低浊度测量，最好使用一个单指数样品池或流动池，这有助于确保测量的足够精度和可重复性。使用无颗粒水，在样品池支架中找到读数最低的样 ( Email/邮箱： info@yh-tek.c o m Phone/电话： 0 21-58220558 W e bsite/网址： w ww.yh-tek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) 品池方向。对于较高的浊度值，可使用不同的样品池。然而，样品池必须匹配(两个不同样品池在标标样品浓度下制备的标准品读数差异必须在±0.005NTU范围内或低于测量精度要求，以较低者为准)。 SAMPLE PREPARATION 样品准备 Prepare the samples as prescribed in the individual monograph. Carefully mix thesamples thoroughly by swirling or inverting the volumetric flask slowly severa times.Avoid shaking or stirring since it may introduce bubbles. Degassing the samples helpsto improve the measurements. For degassing， the samples could stand for severalminutes or a vacuum could be applied， or they could be gently sonicated using anultrasonic bath. After degassing， let the samples stand for several minutes and mix againby carefully inverting two to three times. Transfer the sample to the sample cell andtake the readings. 按照各专题中的规定制备样品。通过缓慢旋转或倒置容量瓶数次，仔细混合样品。避免摇晃或搅拌，因为这可能会产生气泡。对样品进行脱气有助于改进测量。对于脱气，样品可以静置几分钟，或者可以施加真空，或者可以使用超声波浴对其进行轻轻的超声波处理。脱气后，让样品静置几分钟，然后小心地反转两到三次，再次混合。将样品转移至样品池并读取读数。 USE OF FLOW CELLS 流动池的使用 Flow cells are mainly used for low turbidity measurements for samples with smallparticles. When such cells are used， the sample is introduced by carefully pouring itdown the interior edge of the inlet reservoir. In practice， it is advisable to ensure that settling of the particles being measured isnegligible. This is usually accomplished by including a protective colloid in the liquid-suspending medium. It is important that results be interpreted by a comparison ofreadings with those representing known concentrations of suspended matter， producedunder precisely the same conditions. 流动池主要用于小颗粒样品的低浊度测量。当使用这种样品池时，通过小心地将样品倒入进水仓的内边缘来引入样品。 在实际过程中，建议确保被测颗粒的沉降可以忽略不计。这通常通过在液体悬浮介质中加入保护胶体来实现。重要的是，通过将读数与在完全相同的条件下产生的已知悬浮物浓度的读数进行比较来解释结果。 7.2 Nephelometric Procedures 散射光浊度法步骤 Nephelometric procedures are performed similarly to turbidimetric procedures forboth direct measurements and measurements in the ratio mode as described above. 散射光浊度法步骤的执行方式与透射光比浊法程序类似，适用于直接测量和上述比率模式下的测量。 RATE NEPHELOMETRIC PROCEDURES比率模式散射光浊度法步骤 The overall procedure for monitoring the progress of the reaction consists of threewell-defined steps： 1) record a baseline reading of the turbidity of the medium (blank)；2) record the turbidity after the first reagent (antigen) is added， which results in an increase of the turbidity until a plateau is reached； and 3) add the second reagent(antibody)， which results in another turbidity increase and a second plateau followedby a final turbidity increase that continues until a third plateau is reached. Themeasurement zone is selected from the addition of the antibody until the third plateau，depending on the purpose of the assay and the respective component concentrations.Kinetic nephelometry and Endpoint nephelometry are two general procedures that areused for quantifying the immune complexes formed in the immunoassay methods (alsoknownas； immunonephelometry because the measured turbidity isduetoimmunocomplexes that are formed). For each procedure， there are several parametersthat need to be optimized in each individual application. The main parameters are 1)with or without particle enhancement； 2) particle types， sizes， and respective optimumwavelength， if applicable； 3)) monitoringreaction kinetics or endpoint；4)antibody/antigen under consideration and， related to that， the optimum level of antigenloading； 5) buffers and other ionic species and respective optimal pH； 6) type andconcentration of polymers used to modify the solubility of proteins； and 7) temperatureand other environmental factors. Generally these parameters are optimized during themethod development and the values are given in specific monograph(s)and/or chapter(s)as applicable. 监测反应进程的总体程序包括三个明确定义的步骤：1)记录介质浊度的基线读数(空白)；2)在添加一种试剂(抗原)后，记录浊度，这会导致浊度增加，直到达到一个稳定期；3)添加第二种试剂(抗体)，这会导致另一个浊度增加和第二个稳定期，然后是最终浊度增加，直到达到第三个稳定器。根据分析目的和各自的组分浓度，从添加抗体到第三个稳定期中间选择测量区。动力学散射比浊法和终点散射比浊法是两种通用程序，用于量化免疫分析方法中形成的免疫复合物(也称为免疫散射比浊法，因为测得的浊度是由形成的免疫复合物引起的)。对于每一个步骤，都有几个参数需要在每个单独的应用中进行优化。主要参数为1)有无粒子增强；2)颗粒类型、尺寸和各自的最佳波长(如适用)；13监)测反应动力学或终点；4)考虑中的抗体/抗原，以及与之相关的抗原负载的最佳水平；5)缓冲液和其他离子种类以及各自的最佳pH值；6)用于改变蛋白质溶解度的聚合物的类型和浓度；7)温度和其他环境因素。通常，这些参数在方法开发过程中进行了优化，具体的专著和/或章节(如适用)中给出了这些值。 Kinetic nephelometry： The kinetic nephelometry is advantageous compared to theendpoint nephelometry mainly because of the capability to take a sample blank readingin addition to a reagent blank reading. This procedure assesses the rate of theimmunocomplex formation based on the increased intensity response of the scatteredlight of the chosen wavelength. The reaction kinetic may be monitored continuously ora certain number of data points may be taken， depending on the time response of theinstrument used and the type of application. At times it may involve only two datapoints； however， proper care must be exercised because the choice of point selectioncan influence the overall accuracy in cases where differences in reaction kinetics existbetween samples and calibrating standards. Careful consideration should be given tothe appropriate choice of specificity control strategy. 动力学散射比浊法：与终点散射比浊法相比，动力学散射比浊法具有优势，主要是因为除了试剂空白读数外，还能够读取样品空白读数。该程序基于所选波长的散射光的增强强度响应来评估免疫复合物的形成速率。根据所用仪器的时间响应和应用类型，可连续监测反应动力学，或采集一定数量的数据点。有时它可能只涉及两个数据点；但是在样品和校准标准之间存在反应动力学差异的情况下，选择点可能会影响整体准确度。应仔细考虑特异性控制策略的适当选择。 Endpoint nephelometry： In this method， an initial measurement is performedbefore adding the reagent， which represents the blank reading. A second measurementis performed after the immune complex is formed after approximately 60 min. Thedifference between these two measurements is proportional to the content of thecomponent being assayed. 终点散射比浊法：在该方法中，在添加试剂之前进行初始测量，这代表空白读数。大约60分钟后，在免疫复合物形成后进行第二次测量。这两次测量之间的差异与所分析成分的含量成正比。 8.VALIDATION AND VERIFICATION验证与核查 1.Validation 验证 Validation is required when a nephelometric/turbidimetric method is intended foruse as an alternative to the official procedure for testing an official article. The objectiveof nephelometric/turbidimetric method1validationis； to demonstrate that themeasurement is suitable for its intended purpose， including quantitative determinationof the main component in a drug substance or a drug product (Category I assays)，quantitative determination of impurities or limit tests (Category II)， and identificationtests (Category IV). Depending on the category of the test (see Validation ofCompendial Procedures <1225>， Table 2)， the analytical method validation process fornephelometry/turbidimetry requires testing for accuracy， precision， specificity，detection limit (DL)， quantitation limit (QL)， linearity， range， and robustness. Theseanalytical performance characteristics apply to externally standardized procedures andthose that use standard additions. 当散射比浊法/透射浊度法拟用作官方物品测试程序的替代方法时，需要进行验证。当散射比浊法/透射浊度法验证的目的是证明测量适用于其预期目的，包括原料药或药品中主要成分的定量测定(I类分析)、杂质的定量测定或限度试验(Ⅱ类)以及鉴定试验(IV类)。根据试验的类别(参见<1225>Validationof Compendial Procedures， 表2)，透射浊度法/散射比浊法的分析方法验证过程需要对准确度、精密度、特异性、检测限(DL)、定量限(QL)、线性、范围和稳健性进行试验。这些分析性能特征适用于外部标准化程序和那些使用标准添加的程序。 Validation of Compendial Procedures <1225> provides definitions and generalguidance on analytical procedures validation without indicating specific validationcriteria for each characteristic. The intention of the following sections is to provide theuser with specific validation criteria that represent the minimum expectations for this ( Email/邮箱： info@ y h-tek.co m Phone/电话： 0 21-58220558 W V ebsite/网址： w ww .y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) technology. For each particular application， tighter criteria may be needed in order todemonstrate suitability for the intended use. <1225>Validation of Compendial Procedures 章节提供了分析程序验证的定义和一般指南，但没有说明每个特征的具体验证标准。以下各节的目的是向用户提供具体的验证标准，这些标准代表了对该技术的最低期望。对于每个特定应用，可能需要更严格的标准，以证明其适用于预期用途。 ACCURACY 准确度 For Category I， II， and III procedures， accuracy can be determined by conductingrecovery studies with the appropriate matrix spiked with known concentrations of theanalyte. Analysts can also compare the assay results obtained using the nephelometric/turbidimetric procedure under validation to those from an established analyticalprocedure. Validation criteria： 98.0%-102.0% mean recovery for the drug substances， 95.0%-105.0% mean recovery for the drug product assay， and 80.0%-120.0% mean recoveryfor the impurity analysis. These criteria are met throughout the specified range. 对于Ⅰ类、Ⅱ类和ⅢI类程序，可通过使用加入已知分析物浓度的适当基质进行回收研究来确定准确度。分析员还可以将使用验证中的散射光浊度法/透射光比浊法程序获得的分析结果与已建立的分析程序获得的结果进行比较。 验证标准：原料药的平均回收率为98.0%-102.0%，药品分析的平均回收率为95.0%-105.0%，杂质分析的平均回收率为 80.0%-120.0%。这些标准在整个规定范围内都得到满足。 PRECISION精度 Repeatability： The repeatability of the analytical procedure is assessed by measuringthe concentrations of six independently prepared sample solutions at 100% of the assaytest concentration. Alternatively， it can be assessed by measuring the concentrations ofthree replicates of three separate sample solutions at different concentrations. The threeconcentrations should be close enough so that the repeatability is constant across theconcentration range. If this is done， the repeatability at the three concentrations ispooled for comparison to the acceptance criteria. Validation criteria： The relative standard deviation is NMT 1.0% for the drugsubstance， NMT 2.0% for the drug product assay， and NMT 20.0% for the impurityanalysis. 重复性：通过测量六种独立制备的样品溶液在100%分析试验浓度下的浓度来评估分析程序的重复性。或者，可以通过测量三种不同浓度的单独样品溶液的三个重复的浓度来评估。三种浓度应足够接近，以便在整个浓度范围内重复性保持恒定。如果这样做，将三种浓度下的重复性汇总，以与验收标准进行比较。 验证标准：原料药的相对标准偏差为 NMT 1.0%，药品分析的相对标准偏差为NMT2.0%， 杂质分析的相对标准偏差为 NMT 20.0%。 Intermediate precision： The effect of random events on the analytical precision of themethod must be established. Typical variables include performing the analysis ondifferent days， using different instrumentation， and/or having the method performed by two or more analysts. At a minimum， any combination of at least two of these factorstotaling six experiments will provide an estimation of intermediate precision. Validation criteria： The relative standard deviation is NMT 1.5% for the drugsubstance， NMT 3.0% for the drug product assay， and NMT 25.0% for the impurityanalysis. 中间精度： 必须确定随机事件对方法分析精度的影响。典型的变量包括在不同的日期使用不同的仪器进行分析，和/或由两名或两名以上的分析员进行分析。至少，这些因素中的至少两个的组合，总共6个实验，将提供中等精度的评评。 验证标准：原料药的相对标准偏差为 NMT1.5%，药品分析的相对标准偏差为 NMT 3.0%，杂质分析的相对标准偏差为 NMT 25.0%。 SPECIFICITY 特异性 In nephelometric/turbidimetric measurements， specificity is demonstrated by thelack of interference from other components present in the matrix (other components ofthe matrix produce a true solution). 在散射光浊度法/透射光比浊法的浊度测量中，特异性通过基质中其他成分的干扰(基质的其他成分产生真实溶液)的缺乏来证明。 DETECTION LIMIT 检测限 The DL can be estimated by calculating the concentration of a solution that wouldgive the signal-to-noise ratio of ≥ 3.3. The estimated DL must be confirmed byanalyzing samples at the calculated concentration. 可以通过计算溶液的浓度来估计检测限 DL， 该浓度将给出信号的信噪比≥3.3.必须通过分析计算浓度下的样品来确认估计的DL。 QUANTITATION LIMIT 定量限 The QL can be estimated by calculating the concentration of a solution that wouldgive the signal-to-noise ratio of ≥10.0. The estimated QL must be confirmed byanalyzing samples at the calculated concentration. Measurement of a test solutionprepared from a representative sample matrix spiked at the required QL concentrationmust be performed to confirm sufficient sensitivity and adequate precision. Theobserved signal-to-noise ratio at the required QL should be>10. Validation criteria： For the estimated limit of quantitation to be considered valid， themeasured concentration must be accurate and precise at a level ≤50% of thespecification. 定量限 QL 可以通过计算溶液的浓度来估算，该浓度将给出信号的信噪比≥10.0.必须通过分析计算浓度下的样品来确认估算的QL。必须对以所需 QL浓度添加的代表性样品基质制备的试液进行测量，以确认其具有足够的灵敏度和精度。在所需 QL 下观察到的信噪比应大于10. 验证标准：估计的定量限被认为是有效的，测量的浓度必须是准确的，并且在≤50%的规格水平上是精确的。 LINEARITY 线性 A linear relationship between the analyte concentration and measured turbidity response must be demonstrated by preparation of at least four standard solutions atconcentrations encompassing the anticipated concentration of the test solution. Thestandard curve is then evaluated using appropriate statistical methods such as a least-squares regression. Deviation from linearity results from instrumental or sample factors，or both， can be reduced to acceptable levels by reducing or increasing the analyteconcentration， thereby respectively decreasing or increasing the turbidity readings towithin the nephelometer/turbidimeter instrument linearity range. Validation criteria： The correlation coefficient (R) must be NLT 0.995 for Category Iassays and NLT 0.99 for Category II quantitative tests. 分析物浓度和测得的浊度响应之间的线性关系必须通过制备至少四种标准溶液来证明，其浓度包括试验溶液的预期浓度。然后使用适当的统计方法(如最/小二乘回归)评估标准曲线。通过降低或增加分析物浓度，可将仪器或样品因素或两者的线性偏差降低至可接受水平，从而分别将浊度读数降低或增加至透射光法浊度计/散射光浊度计仪器线性范围内。 验证标准：对于Ⅰ类分析，相关系数(R)必须为 NLT0.995， 对于II类定量测试，相关系数(R)必须为 NLT0.99。 RANGE 范围 The operational range of an analytical instrument (and the analytical procedure asa whole) is the interval between the upper and lower concentrations (amounts) ofanalyte in the sample (including these concentrations) for which it has beendemonstrated that the instrumental response function has a suitable level of precision，accuracy， and linearity. Validation criteria： For Category I tests， the validation range for 100.0% centered acceptance criteria is 80.0%-120.0%. For non-centered acceptance criteria， thevalidation range is 10.0% below the lower limit to 10.0% above the upper limit. ForCategory II tests， the validation range covers 50.0%-120.0% of the acceptance criteria. 分析仪器(以及整个分析程序)的操作范围是样品中分析物的上下浓度(数量)(包括这些浓度)之间的间隔，已证明仪器响应函数具有适当的精度、准确度和线性水平。 验证标准：对于I类试验，100.0%中心验收标准的验证范围为 80.0%-120.0%。对于非中心验收标准，验证范围为下限以下10.0%到上限以上10.0%。对于Ⅱ类试验，验证范围涵盖验收标准的50.0%-120.0%。 ROBUSTNESS稳健性 The reliability of an analytical measurement is demonstrated by deliberate changesto experimental parameters. For nephelometry/turbidimetry this can include， forexample， measuring the stability of the analyte under specified storage conditions，varying pH， and adding possible interfering species. RobustnessSis determinedconcurrently using a suitable design for the experimental procedure. 分析测量的可靠性通过有意改变实验参数来证明。对于散射光浊度法/透射光比浊法，这可以包括：测量分析物在特定储存条件、变化的 pH 值和添加可能的干扰物质下的稳定性。使用适合实验程序的设计，同时确保稳健性。 ( Email/邮箱： inf o @yh-tek.co m Phone/电话：021-58220558Website/网址： w ww .y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) 8.2Verification 核查 Current U.S. Good Manufacturing Practices regulations [21 CFR 211.194(a)(2)]indicate that users of analytical procedures described in the U.S. Pharmacopeia andNational Formulary are not required to validate these procedures if provided in amonograph. Instead， they simply must verify their suitability under actual conditions ofuse. 现行的《美国生产规范条例》[21 CFR 211.194(a)(2)表明，如果专论中提供了这些程序，则美国药典和国家处方集中描述的分析程序的用户无需验证这些程序。相反，他们只需验证其在实际使用条件下的适用性。 The objective of nephelometric/turbidimetric procedure verification iis todemonstrate the suitability of a test procedure under actual conditions of use.Performance characteristics that verify the suitability of a nephelometric/turbidimetricprocedure are similar to those required for any analytical procedure. A discussion ofthe applicable general principles is found in Verification of Compendial Procedures<1226>. Verification is usually performed using a reference material and a well-definedmatrix. Verification of compendial nephelometric/turbidimetric procedures includes， atminimum， the execution of the validation parameters for specificity，accuracy，precision， and QL， when appropriate， as indicated in 8.1 Validation. 散射光浊度法/透射光比浊法程序验证的目的是证明测试程序在实际使用条件下的适用性。验证散射光浊度法/透射光比浊法程序适用性的性能特征与任何分析程序所需的性能特征相似。适用的一般原则的讨论见<1226>Verification ofCompendial Procedure 章节。通常使用参考材料和明确定义的基质进行验证。药典散射光浊度法/透射光比浊法程序的验证至少包括对特异性、准确度、精密度和 QL的验证参数的执行(如8.1验证中所述)。 欧洲药典EP10.0 2.2.1. CLARITY AND DEGREE OF OPALESCENCE OFLIQUIDS 液体的澄清度和乳光度 Opalescence is the effect of light being absorbed or scattered by submicroscopicparticles or optical density inhomogeneities. The absence of any particles orinhomogeneities in a solution results in a clear solution. 光被亚微观粒子吸收或散射、或光密度不均匀的产生的效果即为乳光。溶液中不存在任何粒子或不均匀性，就会得到清澈的溶液。 A liquid is considered clear if its clarity is the same as that of water R or of thesolvent used， or if its opalescence is not more pronounced than that of referencesuspension I (see Table 2.2.1.-1)，when examined under the conditions described below. 在下述条件下检查时，如果液体的透明度与水或所用溶剂的透明度相同，或者其乳光不比参考悬浮液Ⅰ(见表2.2.1.-1)的乳光更明显，则认为液体是透明的。 Requirements in monographs are expressed in terms of the visual method bycomparing with the defined reference suspensions (see Table 2.2.1.-1). However，instrumental methods may also be used for determining compliance with monographrequirements once the suitability of the instrument has been established as described belowand calibration with reference suspensions I-IV and with water R or the solvent used hasbeen performed. 通过与规定的参考悬浮液进行比较(见表2.2.1.-1)，以目视法表达专著中的要求。然而，一旦仪器的适用性如下所述建立，仪器方法也可用于确定是否符合专论要求，并使用参考悬浮液 I-IV 和水或所用溶剂进行校准。 VISUAL METHOD 目视法 Using identical test-tubes of colourless， transparent， neutral glass with a flat baseand an internal diameter of 15-25 mm， compare the liquid to be examined with areference suspension freshly prepared as described below. Ensure that the depths ofthelayers in the 2 test-tubes are the same (about 40 mm). 使用相同的无色透明中性玻璃试管，底座平坦，内径为15-25mm，将待检液体与下述新制备的参考悬浮液进行比较。确保两个试管中各层的深度相同(约40mm) o Compare the liquids in diffused daylight 5 min after preparation of the referencesuspension， viewing vertically against a black background. 制备参考悬浮液5分钟后，在漫射日光下比较液体，在黑色背景下垂直观察。 System suitability. The diffusion of light must be such that reference suspension I canreadily be distinguished from water R， and that reference suspension II can readily bedistinguished from reference suspension I (see Table 2.2.1.-1). 系统适用性。光的扩散必须确保参考悬浮液Ⅰ可以很容易地与水区分开，并且参考悬浮液Ⅱ可以很容易地与参考悬浮液Ⅰ区分开(见表2.2.1.-1)。 INSTRUMENTALMETHOD 仪器法 The instrumental assessment of clarity and opalescence provides；amorediscriminatory test that does not depend on the visual acuity of the analyst. Numericalresults are more useful for process control and quality monitoring， especially in stabilitystudies. For example， previous numerical data on stability can be extrapolated todetermine whether a given batch of a preparation will exceed shelf-life limits prior tothe expiry date. 仪器法评估给透明度和乳光度的提供了一种更具辨别力的测试，它不依赖于分析人员的视力。数值结果对于过程控制和质量监控更有用，尤其是在稳定性研究中。 ( Email/邮箱： info@ y h-tek.co m Phone/电话： 0 21-58220558 W V ebsite/网址： w ww.y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) 例如，可以从以前关于稳定性的数字数据外推，来确定给定批次的制剂是否会在有效期之前超过保质期限制。 TURBIDIMETRY AND NEPHELOMETRY 比浊法和浊度法 When a suspension is viewed at right angles to the direction of the incident light，the system appears opalescent due to the scattering of light by the particles of thesuspension (Tyndall effect). A certain portion of the light beam entering a turbid liquidis transmitted， another portion is absorbed and the remaining portion is scattered by thesuspended particles. The light-scattering effect of suspended particles can be measuredeither indirectly by observation of the transmitted light (turbidimetry) or directly bymeasuring the scattered light (nephelometry). Turbidimetry and nephelometry are morereliable in low turbidity ranges， where there is a linear relationship between turbidityvalues and detector signals. As the degree ofturbidity increases， not all the particles areexposed to the incident light and the scattered or the transmitted radiation of otherparticles is hindered on its way to the detector. 当以与入射光方向成直角的角度观察悬浮液时，由于悬浮液颗粒对光的散射(丁达尔效应)，系统呈现乳白色。进入混浊液体的光束的一部分被透射，另一部分被 吸收，其余部分被悬浮颗粒散射。悬浮颗粒的光散射效应可以通过观察透射光(比浊法)间接测量，也可以通过测量散射光(浊度法)直接测量。比浊法和浊度法在低浊度范围内更可靠，浊度值和检测器信号之间存在线性关系。随着浊度的增加， 并非所有粒子都暴露在入射光下，其他粒子的散射或透射辐射在到达探测器的过程中会受到阻碍。 For quantitative measurements， the construction of calibration curves is essential.Linearity must be based on at least 4 levels of concentrations. Reference suspensions mustshow a sufficiently stable degree of turbidity and must be produced under well-definedconditions. 对于定量测量，校准曲线的构建至关重要。线性必须基于至少4个浓度水平。参考悬浮液必须显示足够稳定的浊度，并且必须在明确的条件下产生。 MEASUREMENTS IN RATIO MODE比率模式下的测量 The determination of opalescence of coloured liquids is done using instruments with ratiomode， since colour provides a negative interference， attenuating both incident and scatteredlight and lowering the turbidity value. The effect is so great， even for moderately colouredsamples， that conventional nephelometers cannot be used. 由于颜色会产生负干扰，衰减入射光和散射光，降低浊度值，因此使用具有比率模式的仪器测定有色液体的乳光。这种影响是如此之大，即使是中等颜色的样品，以至于不能使用传统的浊度计。 In turbidimetry or nephelometry with ratio mode， the ratio of the transmissionmeasurement to the 90° scattered light measurement is determined. This procedurecompensates for the light that is diminished by the colour of the sample. Instruments withratio mode use as light source a tungsten lamp with spectral sensitivity at about 550 nmoperating at a filament colour temperature of 2700 K. Other suitable light sources may also ( Email/邮箱： info@ y h-tek.co m Phone/电话： 0 21-58220558 W V ebsite/网址： w ww .y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) be used. Silicon photodiodes and photomultipliers are commonly used as detectors andrecord changes in light scattered or transmitted by the sample. The light scattered at 90 ±2.5° is measured by the primary detector. Other detectors measure back and forward scatter(reflected light) as well as transmitted light. The results are obtained by calculating the ratioof the 90°scattered light measured to the sum of the components of forward scattered andtransmitted light values. 在比浊法或浊度法中，通过比率模式，确定透射测量与90°散射光测量的比率。该程序补偿因样品颜色而减弱的光线。具有比率模式的仪器使用光谱灵敏度约为550 nm 的钨灯作为光源，在2700K的灯丝色温下工作。也可以使用其他合适的光源。硅光电二极管和光电倍增管常用作探测器，记录样品散射或透射光的变化。由主探测器测量90±2.5°处的散射光。其他探测器测量前后散射(反射光)以及透射光。通过计算测得的90°散射光与前向散射光和透射光值分量之和的比值，可以获得结果。 The instruments used are calibrated against standards of known turbidity and arecapable of automatic measurement of turbidity. The test results are obtained directly fromthe instrument and compared to the specifications in the individual monograph. 使用的仪器根据已知浊度标准进行校准，并能够自动测量浊度。测试结果直接从仪器中获得，并与各专著中的规范进行比较。 Alternatively， the influence of the colour of the sample may also be eliminated byusing an infrared light-emitting diode (IR LED) having an emission maximum at 860 nmwith a 60 nm spectral bandwidth as the light source of the instrument. 或者，也可以通过使用最大发射波长为860nm、光谱带宽为 60nm 的红外发光二极管(IRLED)作为仪器光源来消除样品颜色的影响。 INSTRUMENT REQUIREMENTS 仪器要求 Instruments complying with the following characteristics and verified usingreference suspensions as described below may be used instead of visual examinationfor determination of compliance with monograph requirements. 可使用符合以下特征并使用下述参考悬浮液验证的仪器代替目视检查，以确定是否符合专论要求。 -Measuring unit： NTU (nephelometric turbidity units). NTU is based on the turbidity of aprimary standard of formazin. FTU (formazin turbidity units) or FNU (formazinnephelometric units) are also used， and are equivalent to NTU in regions of low turbidity(up to 40 NTU). These units are used in all 3 instrumental methods (nephelometry，turbidimetry and in ratio mode). Measuring range： 0.01-1100 NTU. Resolution： 0.01 NTU within the range 0-9.99 NTU； 0.1 NTU within the range 10.0- 99.9NTU； and 1 NTU for the range> 100 NTU. - Accuracy：±(10 per cent of reading + 0.01 NTU) with in the range 0-20 NTU；± 7.5per cent within the range 20-1100 NTU. -Repeatability： ±0.05 NTU within the range0-20 NTU；±2 per cent of the reading withinthe range 20-1100 NTU. 测量单位：NTU(浊度测量单位)。NTU 是基于福尔马肼一级标准品的浊度。也可使用 FTU(福尔马肼浊度单位)或 FNU(福尔马肼浊度单位)，相当于NTU 的在低浊度区域(最高40NTU)。这些单位适用于所有3种仪器方法(比浊法、浊度法和比率模式)。 -测量范围：0.01-1100 NTU -分辨率： 0-9.99NTU范围内为 0.01 NTU； 10.0-99.9NTU范围内为0.1NTU； 对于大于100 NTU的范围，则为1NTU -准确度：范围在 0-20 NTU 之间，读数准确度偏差为±(读数的10%+0.01NTU)；范围在20-1100 NTU时， 读数准确偏差为±7.5%。 -重复性：在0-20NTU 范围内重复性为±0.05 NTU； 在 20-1100 NTU 范围内读数重复性为±2%。 Instruments with measuring range or resolution， accuracy and repeatabilitycapabilities other than those mentioned above may be used provided they aresufficiently validated and are capable for the intended use. 测量范围或分辨率、精度和重复性能力不同于上述测量范围或分辨率、精度和重复性能力的仪器经过有效验证，也能够应用于预期用途。 CONTROL OF INSTRUMENT PERFORMANCE仪器性能的控制 -Calibration： performed with at least 4 reference suspensions of formazin covering themeasuring range of interest. Reference suspensions described in this chapter or suitablereference standards calibrated against the primary reference suspensions may be used. -校准：使用至少4种福尔马肼参考悬浮液进行校准，覆盖感兴趣的测量范围。可使用本章所述的参考悬浮液或根据主要参考悬浮液校准的适当参考标准。 -Stray light： <0.15 NTU within the range 0-10 NTU；<0.5 NTU within the range 10-1100 NTU. Stray light is defined as that light that reaches the nephelometric detectorwithout being a result of scatter from the sample. Stray light is always a positiveinterference and is a significant source of error in low-range turbidity measurements.Sources of stray light include： imperfections in and scratches on sample cells， internalreflections of the optical system， contamination of the optics or sample cell chamber withdust， and electronic noise. Instrument design can also affect stray light. The influence ofstray light becomes negligible in ratio mode measurements. 杂散光：在 0-10NTU 范围内<0.15 NTU； 在10-1100 NTU 范围内<0.5 NTU。杂散光是指到达浊度检测器的光，而不是样品散射的结果。杂散光总是一种正干扰，是低范围浊度测量中的一个重要误差源。杂散光的来源包括：样品池中的缺陷和划痕、光学系统的内部反射、光学元件或样品池被灰尘污染，以及电子噪声。仪器设计也会影响杂散光。在比率模式测量中，杂散光的影响可以忽略不计。 The test methodology for the specific substance/product to be analysed must also beverified to demonstrate its analytical capability. The instrument and methodology shall beconsistent with the attributes of the substance to be examined. 还必须验证待分析特定物质/产品的试验方法，以证明其分析能力。仪器和方法应与待检物质的属性一致。 Measurements of standards and samples should be carried out under the sametemperature conditions， preferably between 20 °C and 25°℃. 标准品和样品的测量应在相同的温度条件下进行，最好在20°℃和25℃之间。 REFERENCE SUSPENSIONS 参考悬浮液 Formazin has several desirable characteristics that make it an excellent turbidity standard.It can be reproducibly prepared from assayed raw materials. The physical characteristicsmake it a desirable light-scatter calibration standard. The formazin polymer consists ofchains of different lengths， which fold into random configurations. This results in a widevariety of particle shapes and sizes， which allows the analysis of different particle sizes andshapes that are found in real samples. Stabilised formazin suspensions that can be used toprepare stable， diluted turbidity standards are commercially available and may be used aftercomparison with the standards prepared as described. 福尔马肼有几个理想的特性，使其成为一个优秀的浊度液标准。它可以从经过分析的原材料中重复制备。其物理特性使其成为理想的光散射校准标准。福尔马肼聚合物由不同长度的链组成，这些链折叠成随机构型。这会产生各种各样形状和尺寸的颗粒，从而可以分析真实样品中发现的不同颗粒大小和形状。可用于制备稳定稀释浊度标准品的稳定福尔马肼悬浮液是可商购的，并可在与所述制备的标准品进行比较后使用。 All steps of the preparation of reference suspensions as described below are carriedout at 25±3°C. 下述参考悬浮液制备的所有步骤均在25±3°C下进行。 Hydrazine sulfate solution. Dissolve 1.0 g of hydrazine sulfate R in water R anddilute to 100.0 mL with the same solvent. Allow to stand for 4-6 h. 硫酸肼溶液。将1.0g硫酸肼溶解在水中，并用相同的溶剂稀释至100.0mL.静置4-6小时。 Primary opalescent suspension (formazin suspension). In a 100 mL ground-glass-stoppered flask， dissolve 2.5g of hexamethylenetetramine R in 25.0 mL of water R. Add25.0 mL ofthe hydrazine sulfate solution. Mix and allow to stand for 24 h. This suspensionis stable for2 months， provided it is stored in a glass container free from surface defects.The suspension must not adhere to the glass and must be mixed thoroughly before use. 初级乳白色悬浮液(福尔马尔悬浮液)。在100 mL 磨砂玻璃塞烧瓶中，将2.5g六亚甲基四胺溶解在25.0 mL 水中。添加25.0 mL 硫酸肼溶液。混合并静置24小时。如果该悬浮液储存在无表面缺陷的玻璃容器中，则可稳定2个月。悬浮液不得粘附在玻璃上，使用前必须彻底混合。 Standard of opalescence. Dilute 15.0 mL of the primary opalescent suspension to1000.0 mL with water R. This suspension is freshly prepared and may be stored for up to24 h. 乳白色的标准浊度液。用水将15.0 mL 初级乳白色悬浮液稀释至1000.0mL。该悬浮液是新制备的，可储存24小时。 Reference suspensions. Prepare the reference suspensions according to Table 2.2.1.-1. Mix and shake before use. 参考悬浮液。根据表2.2.1-1制备参考悬浮液。使用前混合并摇匀。 Table 2.2.1.-1 I II III IV Standard of opalescence 5.0 mL 10.0 mL 30.0 mL 50.0 mL Water R 95.0 mL 90.0 mL 70.0 mL 50.0 mL Measurements of reference suspensions I-IV in ratio mode show a linear relationshipbetween the concentrations and measured NTU values (see Table 2.2.1.-2). 在比率模式下，参考悬浮液 I-IV的测量结果显示，浓度与测量的 NTU 值之间存在线性关系(见表2.2.1.-2)。 Table 2.2.1.-2 Formazin suspensions Opalescent values (NTU) Reference suspension I 3 Reference suspension II 6 Reference suspension III 18 Reference suspension IV 30 Standard of opalescence 60 Primary opalescent suspension 4000 日本药典17版 2.61 Turbidity Measurement 浊度测量 Turbidity measurement is used to determine the turbidity (degree of opalescence)for the decision whether the article to be examined complies with the clarityrequirement stated in the Purity. As a rule， the visual method is specified for the requirement in individualmonograph. 浊度测量用于确定浊度(乳光度)，以决定待检查的物品是否符合纯度中规定的透明度要求。 作为一项规则，，目视法是针对个别专论中的要求说明的。 1. Visual method目视法 This is used to determine the degree of opalescence with white (or faintly-colored)fine particles. So the degree of opalescence of a colored sample is liable to bedetermined lower that it is difficult to compare the degree correctly without usingsimilarly colored reference suspension. 这是用来确定乳白色(或淡色)细颗粒的乳光程度。因此，有色样品的乳光度容易被测定得较低，因此，如果不使用类似颜色的参考悬浮液，就很难正确地比较其乳光度。 1. Reference suspensions 参考悬浮液 Pipet 5 mL，10 mL， 30 mL and 50 mL of formazin opalescence standard solution，dilute them separately to exactly 100 mL with water， and use these solutions so obtainedas Reference suspensions I， II， III and IV， respectively. Shake before use. Degrees ofopalescence of Reference suspensions I， II， III and IV are equivalent to 3 NTU， 6 NTU，18 NTU and 30 NTU， respectively. 用移液管分别吸取5 mL、10 mL、30 mL、50mL福尔马肼标准液，用水分别稀释至100mL，分别作为参比悬液Ⅰ、II、ⅢI、ⅣV。在使用前摇晃。参考悬浮液I、I、Ⅲ和ⅣV的乳光度分别相当于3NTU、6NTU、18NTU 和 30 NTU. 2. Procedure步骤 Place sufficient of the test solution， water or the solvent to prepare the test solutionand， where necessary， newly prepared Reference suspensions in separate flat-bottomedtest tubes， 15 -25 mm in inside diameter and of colorless and transparent， to a depth of40 mm， and compare the contents of the tubes against a black background by viewingin diffused light down the vertical axes of the tubes. The diffused light must be suchthat Reference suspension I can be readily distinguished from water， and that Referencesuspension II can readily be distinguished from Reference suspension I. 取足够的待测溶液、水或溶剂，以准备测试溶液，必要时，将新制备的参考悬浮液置于独独的平底试管中，试管内径15-25mm，无色透明，深度40 mm。然后在一个黑色的背景下通过漫射光下垂直于管轴进行观察，比较管内的内容。漫射光必须能使参考悬浮体I容易与水区分开来，参考悬浮体Ⅱ容易与参考悬浮体I区分开来。 In this test Reference suspensions are used when the clarity of the test solution isobscurely and it is not easy to determine that its degree of opalescence is similar or notsimilar to water or to the solvent used to prepare the test solution. 在此测试中，当测试溶液的透明度模糊不清，并且不容易确定其乳光度与水或与用于制备测试溶液的溶剂是否相似时，使用参考悬浮液。 3. Interpretation注释 A liquid is considered“clear"when its clarity is the same as that of water or of thesolvent used to prepare the liquid or its turbidity is not more pronounced than that ofReference suspension I. If the turbidity of the liquid is more than that of Reference ( Email/邮箱： info@ y h-tek.co m Phone/电话： 0 21-58220558 W V ebsite/网址： w ww .y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) suspension I， consider as follows： When the turbidity is more than that of Referencesuspension I but not more than that of Reference suspension II， express “it is not morethan Reference suspension II". In the same way， when the turbidity is more than that ofReference suspension II but not more than that of Reference suspension III， express "itis not more than Reference suspension III”， and when the turbidity is more than that ofReference suspension III but not more than that of Reference suspension IV， express“it is not more than Reference suspension IV". When the turbidity is more than that ofReference suspension IV， express“it is more than Reference suspension IV”. 当液体的澄清度与水或与用于制备液体的溶剂的澄清度相同或其浊度不比参比悬浮液Ⅰ更明显时，该液体被视为“澄清”。如果液体的浊度大于参考悬浮液I， 考虑如下：当浊度大于参考悬浮液Ⅰ但不超过参考悬浮液Ⅱ时，表示“不超过参考悬浮液 II”。同理，当浊度大于参比悬浊液II但不大于参比悬浊液II时， 表示“不大于参比悬浊液III”，当浊度大于参比悬浊液II时但不超过参考悬浮液IV，表示“不超过参考悬浮液IV”。当浊度大于参考悬浮液IV时，表示“大于参考悬浮液IV”。 1.4. Reagent solutions 试剂溶液 Formazin opalescence standard solution： To exactly 3 mL of formazin stocksuspension add water to make exactly 200 mL. Use within 24 hours after preparation.Shake thoroughly before use. Degrees of opalescence of this standard solution isequivalent to 60 NTU. 福尔马津乳光标准溶液：准确地取 3mL福尔马肼储备悬浮浮，加水至200mL。配制后24小时内使用。使用前彻底摇匀。此标准溶液的乳光度相当于60NTU. 2. Photoelectric photometry 光电光度法 The turbidity can also be estimated by instrumental measurement of the lightabsorbed or scattered on account of submicroscopic optical density inhomogeneities ofopalescent solutions and suspensions. The photoelectric photometry is able to providemore objective determination than the visual method. Though they can determine theturbidity by measuring the scattered or transmitted light， the measuring system and lightsource must be specified in individual test method， and for the comparison of observeddata， the same measuring system and light source should be used. 由于乳光溶液和悬浮液的亚显微光密度不均匀性，还可以通过仪器测量吸收或散射的光来估计浊度。光电光度法比目测法能够提供更客观的测定。 虽然他们可以通过测量散射光或透射光来确定浊度，但必须在单独的测试方法中标明测量系统和光源，并且为了比较观察数据，应使用相同的测量系统和光源。 In each case， the linear relationship between turbidity and concentration must bedemonstrated by constructing a calibration curve using at least 4 concentrations. Forcolored samples， the turbidity value is liable to be estimated lower because ofattenuating both incident and scattered lights due to the absorption by the color， and thetransmission-dispersion method is principally used. ( Email/邮箱： inf o @yh-tek.co m Phone/电话：021-58220558Website/网址： w ww .y h - t ek.com Address/地址：上海市浦东新区航川路18号1号楼203室 ) 在每种情况下，浊度和浓度之间的线性关系必须通过使用至少4种浓度构建校准曲线来证明。对于有颜色的样品，由于颜色的吸收，入射光和散射光都被衰减，浊度值容易被估计得较低，主要采用透射-色散法。 1.Turbidimetry 透射光比浊法 When a light passes through a turbid liquid the transmitted light is decreased byscattering with the particles dispersed in the liquid. A linear relationship is observedbetween turbidity and concentration when the particles with a constant size areuniformly dispersed， the size is small and the suspension is not higher concentration.The turbidity can be measured by Ultraviolet-visual Spectrophotometry <2.24> usingspectrophotometer or photoelectric photometer. The turbidity of the sample in higherconcentration can also be measured， however， it is susceptible to the color of the sample，and the measurement is usually performed at around 660 nm to avoid possibledisturbance occurred from the absorption by the color. 当光通过混浊液体时，透射光通过分散在液体中的颗粒散射而减少。当粒径恒定的颗粒分散均匀、粒径较小且悬浮液浓度不高时，浊度与浓度呈线性关系。浊度可以通过紫外分光光度法<2.24>使用分光光度计或光电光度计进行测量。较高浓度的样品的浊度也可以测量，但它易受样品颜色的影响，通常在660 nm 左右进行测量，以避免颜色吸收可能产生的干扰。 2. Nephelometry 散射光浊度法 When a suspension is viewed at right angles to the direction of the incident light，it appears opalescent due to the refraction of light from the particles of the suspension(Tyndall effect). A certain portion of the light entering a turbid liquid is transmitted，another portion is absorbed and the remaining portion is scattered by the suspendedparticles. The scattered light measuring method shows the linear relationship betweenthe nephelometric turbidity units (NTU) values and relative detector signals in a lowturbidity range. As the degree of turbidity increases， not all the particles are exposed tothe incident light and the scattered radiation of other particles is hindered on its way tothe detector. 当悬浮物与入射光方向成直角时，由于悬浮物粒子的光线折射(丁达尔效应)，悬浮物呈现乳白色。进入混浊液体的光，一部分被透射，一部分被吸收，剩下的部分被悬浮的粒子散射。散射光测量方法显示了低浊度范围内散射浊度单位(NTU)值与相对检测器信号之间的线性关系。随着浊度的增加，并不是所有的粒子都暴露在入射光下，其他粒子的散射辐射在到达探测器的过程中会受到阻碍。 3. Ratio Turbidimetry比率浊度法 This method measures both scattered and transmitted light values at the same time，and the turbidity is determined from the ratio of the scattered light value to thetransmitted light value. This procedure compensates for the light that is diminished bythe color of the sample and eliminates the influence of the color. When the measurementis performed by using an integrating sphere， it is particularly called the integratingsphere method， which measures the total transmitted light value as well as the scattered light value occurred with the suspended particles， and the turbidity can be determinedfrom the ratio of them. 该方法同时测量散射光值和透射光值，浊度由散射光值与透射光值的比值确定。此程序可补偿因样品颜色而减弱的光线，并消除颜色的影响。当用积分球进行测量时，特别称为积分球法，它测量悬浮粒子的总透射光值和散射光值，由它们的比值可以确定浊度。 2.4.Application of photoelectric photometry for monograph requirements 光电光度法在专著要求中的应用 The turbidity of the test solution， determined by the photoelectric photometry， canbe used as an indicating standard for the conformity to the clarity requirements byconverting into NTU by using turbidity known reference solutions such as Referencesuspensions I - IV， if needed， and water or the solvent used. In an automaticallycompensable apparatus being calibrated with turbidity known reference solutions， themeasuring result is given in NTU and it can be compared directly with requiredspecified value. 由光电光度法测定的测试溶液的浊度，可以作为指示标准，通过使用浊度已知的参考溶液，如参考悬浮液I-IV，如果需要，水和使用的溶剂液也可以，将其以 NTU 为单位的数据转出。在使用浊度已知参考溶液校准的自动补偿装置中，测量结果以 NTU 为单位给出，并且可以直接与所需的规定值进行比较。 NTU is often used as the unit in the turbidity determinations. It is the unit used inthe case when the turbidity is estimated by the instrument which measures the 90±30°scattered light against the incident light intensity， using tungsten lamp， and in the casethe estimation is performed by the instrument which measures the 90±2.5°scatteredlight against the incident light intensity using 860 nm infrared light， FNU is used as theunit. FNU is equivalent to NTU at a range of smaller measurements (less than 40 NTU).For the unit of formazin concentration， FTU is also used， which is defined as asuspension of 1 mg formazin in 1L of purified water is 1 FTU. 在浊度测定中经常使用 NTU 作为单位。它是测量用90±30°散射光对入射光强度的得到浊度信息时使用的单位，使用钨灯，采用860 nm 红外光测量90±2.5°的散射光对入射光强度，此时以 FNU 为单元。在较小的测量范围内(小于40NTU)，FNU 相当于 NTU。福尔马肼的浓度单位也用 FTU， 即1L纯净水中1mg福尔马肼的悬浮液为1FTU. Formazin stock suspension. To 25 mL of hexamethylenetetramine TS add 25 mL ofhydrazinium sulfate TS， mix， and use after allowing to stand at room temperature for24 hours. Store in a glass container free from surface defects. Use within 2 months.Shake thoroughly before use. The turbidity of this suspension is equivalent to 4000NTU. 福尔马肼贮备悬浮液：向25mL 六亚甲基四胺中加入25 mL硫酸肼，混匀， 室温静置24小时后使用。 储存在没有表面缺陷的玻璃容器中。2个月内使用。使用前彻底摇匀。这种悬浮液的浊度相当于4000 NTU。 Formazin opalescence standard solution. To 15 mL of formazin stock suspensionadd water to make 1000 mL. Use within 24 hours after preparation. Shake thoroughlybefore use. 福尔马标准液：向15mL的福尔马肼贮备悬浮液中加水至1000mL。配制后24小时内使用。使用前彻底摇匀。 解决方案： 上海胤煌科技针对药剂的澄清度检查推出了以下产品，符合各国药典的溶液澄清度检查规范。 1、澄清度检查专用伞棚灯 HN-100A 型和 HN-200A 型澄清度检查专用伞棚灯符合各国药典中目视法检测溶液澄清度的仪器要求，具有光林带型光源，能有效减少目视过程中光对眼睛的刺激，其照度可达5000 Lux。其中 HN-200A 型专用伞棚灯增加了 RGB 三色光源，可以对有色样品进行澄清度检测。 2、YH-CLS-1201澄清度检查分析仪 此仪器采用全彩液晶触摸屏进行操作控制，可以直接检测注射用原料药和注射剂的澄清度，并具备四级权限管理和审计追踪功能，完全满足 GMP 的数据完整性要求，是液体一致性评价的有效仪器。 Email/邮箱： info@yh-tek.comPhone/电话：ebsite/网址： www.yh-tek.comAddress/地址：上海市浦东新区航川路楼 中/美/欧/日四大药典澄清度检查规范-中英双译中国药典20200902 澄清度检查法澄清度检查法系将药品溶液与规定的浊度标准液相比较，用以检查溶液的澄清度。除另有规定外，应采用第一法进行检测。品种项下规定的“澄清”，系指供试品溶液的澄清度与所用溶剂相同，或不超过0.5号浊度标准。“几乎澄清”，系指供试品溶液的浊度介于0.5号至1号浊度标准液的浊度之间。第一法（目视法）除另有规定外，按各品种项下规定的浓度要求，在室温条件下将用水稀释至一定浓度的供试品溶液与等量的浊度标准液分别置于配对的比浊用玻璃管（内径15-16 mm,平底，具塞，以无色、透明、中性硬质玻璃制成）中，在浊度标准液制备5 分钟后，在暗室内垂直置于伞棚灯下，照度为1000 lx，从水平方向观察、比较。除另有规定外外，供试品溶解后应立即检视。第一法无法准确判定两者的澄清度差异时，改用第二法进行测定，并以其测定结果进行判定。浊度标准存贮液的制备 称取于105℃干燥至恒重的硫酸肼1.00 g，置于100 ml量瓶中，加水适量使溶解，必要时可在40℃的水浴中温热溶解，并用水稀释至刻度，摇匀，放置4-6小时；取此溶液于等容量的10%乌洛托品溶液混合，摇匀，于25℃避光静置24小时，即得。该溶液置冷处避光保存，可在2个月内使用，用前摇匀。浊度标准原液的制备 取浊度标准贮备液15.0 ml，置1000 ml量瓶中，加水稀释至刻度，摇匀，取适量，置1 cm吸收池中，照紫外-可见分光光度法（通则0401），在550 nm的波长处测定，其吸光度在0.12-0.15范围内，该溶液应在48小时内使用，用前摇匀。浊度标准液制备 取浊度标准原液与水，按照下表配置，即得。浊度标准液应临用时制备，使用前充分摇匀。                                               第二法（浊度仪法）供试品的浊度可采用浊度仪测定。溶液中不同大小、不同特性的微粒物质包括有色物质均可使入射光产生散射，通过测定透射光或者散射光的强度，可以检查供试品的浊度。仪器测定模式通常有三种类型，透射光式、散射光式和透射光-散射光比较测量模式（比率浊度模式）。1.仪器的一般要求采用散射光式浊度仪时，光源峰值波长为860 nm；测量范围应包含0.01-100NTU。在0-10NTU范围内分辨率应为0.01NTU；在10-100NTU范围内分辨率应为0.1NTU.2.适用范围及检测原理本法采用散射光式浊度仪，适用于低、中浊度无色供试品溶液的浊度测定（浊度值为100NTU以下的供试品。）因为高浊度的供试品会造成多次散射现象，时散射光强度迅速下降，导致散射光强度不能正确反映供试品的浊度值。0.5-4号浊度标准液的浊度值范围约为0-40NTU。采用散射光式浊度仪测定时，入射光和测定的散射光呈90℃夹角，入射光强度和散射光强度关系式如下。I=K’T I0式中 I为散射光强度，单位为cd；     I0 为入射光强度，单位为cd；     K’为散射系数；     T为供试品溶液的浊度值，单位为NTU（NTU是基于福尔马肼浊度标准液液测定的散射浊度单位，福尔马肼浊度标准液即为第一法中的浊度标准贮备液）。在入射光I0不变的情况下，散射光强度I与浊度值成正比。因此，可以将浊度测量转化为散射光强度的测量。3.系统的适用性试验仪器应定期（一般每月一次）对浊度标准液的线性和重复性进行考察，采用0.5号至4号浊度标准液进行浊度值测定，浊度标准液的测定解果（单位NTU）与浓度间应呈线性关系，线性方程的相关系数应不低于0.999；取0.5号至4号浊度标准液，重复测定5次，0.5号和1号浊度标准液测量浊度值的相对标准偏差应不大于5%，2-4号浊度标准液测量浊度值的相对标准偏差不大于2%。4.测定法按照仪器说明书要求并采用规定的浊度液进行仪器校正。溶液剂直接取样测定；原料药或者其它剂型按照个论项下的标准规定制备供试品溶液，临用时制备。分别取供试品溶液和相应浊度标准液进行测定，测定前应摇匀，并避免产生气泡，读取浊度值。供试品溶液浊度值不得大于相应浊度标准液的浊度值。 美国药典USP44<630> VISUAL COMPARISON 视觉比较The purpose of this test is to provide the details for the visual comparison of the color and/or turbidance of sample solutions of certain concentration to a standard solution or a series of standard solutions of known concentration. Where a color or turbidity comparison is directed, follow the procedures and conditions outlined below for performing these tests.本试验的目的是提供特定浓度的样品溶液与已知浓度的标准溶液或一系列标准溶液的颜色和/或浊度的视觉比较细节。如果需要进行颜色或浊度比较，请遵循以下程序和条件进行这些测试 Comparison vessels: Color-comparison tubes matched as closely as possible in internal diameter, in depth of sample solution, and in all other respects should be used.对比容器：应使用内径、样品溶液深度和所有其他方面尽可能匹配的颜色对比管。 Viewing conditions for turbidity comparison: Tubes should be viewed horizontally against a dark background with the aid of a light source directed from the sides of the tubes.浊度比较的观察条件：应在黑暗背景下，借助从管子侧面发出的光源水平观察管子。 Viewing conditions for color comparison: Tubes should be viewed downward against a white background. Most of the time, common room lighting is sufficient to perform the assessment. A light source directed from beneath the bottoms of the tubes may be used if needed and if the practice is consistent between the materials under comparison.颜色比较的观察条件：管子应在白色背景下向下观察。大多数情况下，公共空间照明足以进行评估。如果需要，并且对比材料之间的实践一致，可以使用从管底部下方引导的光源 <855> NEPHELOMETRY AND TURBIDIMETRY      散射光浊度法和透射光比浊法1. INTRODUCTION 介绍Nephelometry and turbidimetry are analytical techniques that are based on the principles of light-scattering phenomena. Light scattering is the physical phenomenon in which a beam of light changes its direction of propagation (known as deflection) as a result of interaction with sufficiently small matter particles. It has been established from the Maxwell electromagnetic theory that a prerequisite for scattering to occur is that the refractive indexes of the suspended particles must be different from those of the suspending liquid. The larger the difference, the more intense the scattering becomes. There are two types of light scattering: 1) elastic scattering, in which the wavelength of the scattered light and incident light are the same; and 2) inelastic light scattering, in which the wavelength of the scattered light and incident light are different. Only the first type of light scattering (elastic) is relevant to nephelometry and turbidimetry.散射光浊度法和透射光比浊法是基于光散射现象原理的分析技术。光散射是一种物理现象，其中光束由于与足够小的物质粒子相互作用而改变其传播方向（称为偏转）。根据麦克斯韦电磁理论，散射发生的先决条件是悬浮颗粒的折射率必须不同于悬浮液体的折射率。差异越大，散射越强烈。光散射有两种类型：1）弹性散射，其中散射光和入射光的波长相同；2）非弹性光散射，其中散射光和入射光的波长不同。只有前一种光散射（弹性）与散射光浊度法和透射光比浊法有关。 In turbidimetry, the intensity of the transmitted light is measured and the attenuation of the intensity of incident light as a result of scattering is measured at the direction of incident light (i.e., 0°) and compared to the intensity of incident light (blank measurement). The measured property is an indirect measurement of the scattering effect of the suspended particles and is referred to as turbidance. Any absorbance of light by the suspended sample will result in additional attenuation of light intensity (see Ultraviolet-Visible Spectroscopy <857> and Ultraviolet-Visible Spectroscopy—Theory and Practice <1857>). Hence, it is important to ensure that the material being measured does not absorb light at the measurement wavelength. Indeed the equations governing absorption and turbidimetry are the same (albeit with different values for the attenuation constants). In nephelometric techniques, the intensity of the scattered light at a 90° angle from the propagation direction of the incident light is measured. Therefore, a nephelometric measurement is a direct measurement of the scattering effect of suspended matter.在透射光比浊法中，测量透射光的强度，并在入射光方向（即0°）测量散射导致的入射光强度的衰减，并与入射光强度进行比较（空白测量）。被测特性是悬浮颗粒散射效应的间接测量，称为浊度。悬浮样品对光的任何吸收都会导致光强度的额外衰减（参见<857> Ultraviolet-Visible Spectroscopy和<1857> Ultraviolet-Visible Spectroscopy—Theory and Practice）。因此，确保被测材料不会吸收测量波长处的光非常重要。实际上，控制吸收和浊度测定的方程式是相同的（尽管衰减常数的值不同）。在散射光浊度法中，测量与入射光传播方向成90°角的散射光强度。因此，散射光浊度法浊度测量是对悬浮物散射效应的直接测量。 2. TERMS AND DEFINITIONS 术语和定义Terms commonly used in describing turbidimetric and nephelometric techniques are:• Turbidance (symbol, S): A measure of the decrease of the transmitted incident light beam intensity as a result of the light-scattering effect of suspended particles. The amount of suspended matter may be measured by observation of either the transmitted light (turbidimetry) or the scattered light (nephelometry).log I0/It = kbc = TI0 = intensity of incident lightIt = intensity of transmitted lightk = molar turbidity coefficientb = cell path lengthc = concentrationT = turbidance• Turbidity (symbol, τ): In turbidimetric measurements, the turbidity is the measure of the decrease in incident beam intensity/unit length of a given suspension. The International Organization for Standardization defines turbidity as “the reduction of transparency of a liquid caused by the presence of undissolved matter”.• Turbidity Measurement Units: The turbidity units are stated using a descriptor which indicates the method of measurement.• Nephelometric Turbidity Units (NTUs): When the turbidity is measured using a nephelometer, which measures the scattered light at a 90° angle from the direction of propagation of incident light, the units of turbidity are called nephelometric turbidity units (NTUs). The magnitude of NTU is defined based on the turbidity generated by primary formazin standard (a suspension made by mixing solutions of hydrazine sulfate and hexamethylenetetramine in water). Safer polymer-bead suspensions are now commercially available and are recognized as an acceptable alternative. However, all those standards are traced to formazin. The primary formazin standard solution has been assigned a turbidity of 4000 NTUs.Other recognized units for turbidity include the formazin turbidity unit (FTU) and the formazin nephelometric unit (FNU). These units are equivalent to NTU for the range from 0–40 NTUs.描述浊度法和浊度法的常用术语包括：•浊度（符号S）：由于悬浮颗粒的光散射效应，透射入射光束强度降低的一种度量。悬浮物的量可以通过观察透射光（比浊法）或散射光（浊度法）来测量。log I0/It = kbc = TI0=入射光强度It=透射光强度k=摩尔浊度系数b=样品池路径长度c=浓度T=浊度•浊度（符号，τ）：在透射光浊度测量中，浊度是给定悬浮液的入射光束强度/单位长度减少的量度。国际标准化组织将浊度定义为“由于存在未溶解物质而导致液体透明度降低”。•浊度测量单位：浑浊度单位用一个描述符表示，该描述符指示测量方法。•散射光浊度计浊度单位（NTU）：当使用散射光浊度法测量浊度时，浊度计以与入射光传播方向成90°角的角度测量散射光，浊度单位称为散射光浊度法浊度单位（NTU）。NTU的大小是根据初级福尔马肼标准品（一种将硫酸肼和六亚甲基四胺溶液混合在水中制成的悬浮液）产生的浊度定义的。更安全的聚合物微珠悬浮液现已上市，并被公认为可接受的替代品。然而，所有这些标准都可以追溯到福尔马肼。初级福尔马肼标准溶液的浊度为4000 NTU。其他公认的浊度单位包括福尔马肼比浊法单位（FTU）和福尔马肼浊度法单位（FNU）。这些单位相当于0-40 NTU范围内的NTU。3. APPLICATIONS 应用Turbidimetric and nephelometric techniques have applications that include 1) concentration determination of solutions and/or suspensions (determination of several cations and anions by precipitating and suspending the resulting precipitate at well-controlled reaction parameters); 2) measurement of the degree of turbidity of turbid solutions or suspensions; 3) determination of weight-average molecular weights and dimensions of polydisperse systems in the molecular weight range from 1000 to several hundred million; 4) measurement of immunoassays’ reaction kinetics or kinetics of immunoprecipitations (rate nephelometry); 5) monitoring of cell and bacteria growth; and 6) particle size distribution determination of suspended material, particle counting, etc.透射光比浊法和散射光浊度法技术的应用包括1）溶液和/或悬浮液的浓度测定（通过在控制良好的反应参数下沉淀和悬浮产生的沉淀物，来测定几种阳离子和阴离子）；2）测量混浊溶液或悬浮液的浊度；3）测定分子量在1000到数亿之间的多分散体系的重均分子量和尺寸；4）测量免疫分析的反应动力学或免疫沉淀动力学（比率散射浊度法）；5）监测细胞和细菌的生长；6）悬浮物粒度分布测定、颗粒计数等。 Rate nephelometry is widely used for vaccine components assays and/or quantitation of components in blood serum. It is also used for host cell protein qualification in recombinant biopharmaceuticals. When using the technique, the measurement of the change in the light-scattering response by antigen–antiserum or antigen-purified antibody complexes is used to calculate the amount of antigen (Ag) or antibody (Ab) responsible for the immunological Ab-Ag precipitation reaction or agglutination reaction. Often the antigens under consideration are linked covalently or adsorbed to polymeric microspheres to increase the scattering efficiency; the resulting technique is known as "particle-enhanced immunoassay". Although the technique is described as nephelometry, usually both scattered and transmitted light are measured using the ratio instruments.比率散射浊度法广泛用于疫苗成分分析和/或血清成分的定量。它还用于重组生物制药中的宿主细胞蛋白质鉴定。当使用该技术时，通过测量抗原-抗血清或抗原纯化抗体复合物的光散射反应的变化，来计算导致免疫抗体-抗原沉淀反应或凝集反应的抗原（Ag）或抗体（Ab）的量。通常考虑抗原共价连接或吸附在聚合物微球上，以提高散射效率；由此产生的技术被称为“颗粒增强免疫分析”。虽然这项技术被称为散射光浊度法，但通常散射光和透射光都是用比率仪器测量的。 Nephelometric measurements are more reliable in low turbidity ranges (relatively low concentration of the scattering medium). In this range, a linear relationship is observed between the sample concentration and the detector’s signal intensity expressed as NTU. As the concentration increases, so does the incidence of multiple scattering that deviates the response from the linearity. The maximum NTU value, which supports a reliable linearity relationship, is in the range of 1750–2000 NTUs. Turbidimetry is preferred for higher turbidity ranges (concentrations of the scattering media). To achieve consistent results, all measurement variables must be carefully controlled. Where such control is possible, extremely dilute suspensions may be measured.散射光法浊度测量在低浊度范围（散射介质浓度相对较低）更可靠。在该范围内，观察到样品浓度与检测器信号强度（以NTU表示）之间存在线性关系。随着浓度的增加，多次散射的入射角也会增加，从而偏离线性响应。支持可靠线性关系的最大NTU值在1750–2000 NTU范围内。透射光比浊法适用于更高的浊度范围（散射介质的浓度）。为了获得一致的结果，必须仔细控制所有测量变量。在可能的情况下，可以测量极稀的悬浮液。 4. INSTRUMENTATION 仪器仪表Instruments used for turbidimetric and nephelometric measurements are called turbidimeters and nephelometers, respectively. Generally, these instruments consist of a mercury lamp with filters for the strong green or blue lines, a shutter, a set of neutral filters with known transmittance, and a sensitive photomultiplier, which can be mounted fixed at 0° or at a 90° angle from the incident light propagation direction, or on an arm that can be rotated around the solution cell and set at any angle from −135° to 0° to +135° by a dial outside of the light-tight housing. Solution cells are of various shapes, such as square for measuring 90° scattering; semioctagonal for 45°, 90°, and 135° scattering; and cylindrical for scattering at all angles (see Figure 1).用于透射光比浊法和散射光浊度法测量的仪器分别称为透射光浊度计和散射光浊度计。通常，这些仪器包括一个带有滤光器的汞灯（用于强绿线或蓝线）、一个快门、一组具有已知透射率的中性滤光器和一个灵敏的光电倍增管，该光电倍增管可安装在与入射光传播方向成0°或90°角的位置，或者在一个臂上，它可以围绕溶液单元旋转，并通过不透光外壳外的表盘设置为−135°到0°到+135°的任何角度。溶液池的形状多种多样，例如用于测量90°散射的正方形；45°、90°和135°散射为半八角形；圆柱形可适用于所有角度的散射（见图1）。 Figure 1. Representative nephelometric (turbidimetric) instrument. Note that Detector 2 may be mounted on a movable arm.图1。代表性浊度仪。注意，探测器2可安装在可移动臂上。 Turbidity also can be measured with a standard photoelectric filter photometer or spectrophotometer, preferably with illumination in the blue portion of the spectrum. Nephelometric measurements require an instrument with a photocell placed so as to receive scattered, rather than transmitted, light. Because this is the same geometry used in fluorometers, they can be used as nephelometers by proper selection of filters. A ratio turbidimeter combines the technology of 90° nephelometry and turbidimetry. It contains photocells that receive and measure scattered light at a 90° angle from the sample as well as receiving and measuring the forward scatter in front of the sample. It also measures light transmitted directly through the sample. Linearity is attained by calculating the ratio of the 90° angle scattered light measurement to the sum of the forward scattered light measurement and the transmitted light measurement. The benefit of using a ratio turbidimetric system is that the measurement of stray light becomes negligible. In addition, the determination of turbidity of colored suspensions is done exclusively using turbidimetric or nephelometric instruments with ratio mode because this procedure compensates for the attenuation of light as the result of the suspension color. Typically, the light source in these instruments is a tungsten lamp with most of the light intensity at about 550 nm operating at the filament temperature of 2700 K. Other suitable light sources are also available. Typically, the detectors are ▲silicon diodes▲ (ERR 1-May-2019) and photomultipliers. An alternative for eliminating the color effect involves using an infrared light-emitting diode as a light source, which yields an emission maximum centered at about 860 nm and a spectral bandwidth of 60 nm. When laser light sources are also used, especially in nephelometric instruments, the technique is commonly known as "laser nephelometry". The advantage of using laser nephelometers is the significant improvement in signal-to-noise ratio at very low detection levels. Usually the light source is a laser diode with a working wavelength at 660 nm. The high-power density of the laser beam gives rise to higher scattered intensity from smaller particles. Combined with a light trap, which absorbs the unscattered light, the system lowers the stray light significantly. When the use of a nephelometer or turbidimeter is indicated for a procedure in a monograph, instruments working in ratio mode may be used instead.浊度也可以用标准光电滤光光度计或分光光度计测量，最好是在光谱的蓝色部分进行照明。散射光浊度法测量需要一个装有光电管的仪器，以便接收散射光，而不是透射光。由于这与荧光计中使用的几何结构相同，因此可通过适当选择滤光片将其用作浊度计。比率浊度计结合了90°散射光浊度法和透射光比浊法。它包含光电管，接收和测量与样品成90°角的散射光，以及接收和测量样品前面的前向散射光。它还测量直接穿过样品的光。通过90°角散射光测量值，前向散射光测量值和透射光测量值之和，计算两者的比值，可获得线性度。使用比率浊度测量系统的好处是杂散光的测量变得可以忽略不计。此外，彩色悬浮液的浊度测定仅使用透射光比浊法浊度仪或浊度仪（带比率模式）进行，因为该程序补偿了悬浮液颜色导致的光衰减。通常，这些仪器中的光源是钨灯，在2700 K的灯丝温度下工作，大部分光强约为550 nm。也可使用其他合适的光源。通常，探测器是▲硅二极管▲和光电倍增管。另一种消除颜色效应的方法是使用红外发光二极管作为光源，其最大发射中心约为860 nm，光谱带宽为60 nm。当激光光源也被使用时，尤其是在浊度测量仪器中，这种技术通常被称为“激光浊度测量”。使用激光散射光浊度计的优点是，在非常低的检测水平下，信噪比显著提高。通常光源是工作波长为660 nm的激光二极管。激光束的高功率密度使较小粒子产生更高的散射强度。与吸收未散射光的光阱相结合，该系统可显著降低杂散光。当专著中的某个程序指示使用散射光浊度计或透射光浊度计时，可以使用在比率模式下工作的仪器。 5. FORMAZIN TURBIDITY STANDARDS福尔马肼浊度标准Formazin is the only known primary turbidity standard. All other standards are secondary and must be traced to formazin. The primary standard is defined in the ▲IUPAC Compendium of Chemical Terminology,▲ (ERR 1-May-2019) 2nd ed. (the Gold Book) as one that is prepared by the user from traceable materials using well-defined methodologies and conditions.福尔马肼是唯一已知的主要浊度标准。 所有其他标准都是次要的，必须追溯到福尔马肼。 主要标准在▲IUPAC Compendium of Chemical Terminology▲（ERR 1-May-2019）第 2 版（金书）中被定义为由用户使用明确定义的方法和条件从可追溯的材料准备的标准。 Formazin suspension has many features that ensure its suitability as a primary standard. It can be consistently and accurately prepared from reagent-grade starting materials. The suspension consists of random polymers with different lengths and of random configurations, which result in moieties of varying shapes and sizes ranging from less than 0.1 μm to more than 10 μm. Although the polymer chain length distribution has been shown to vary from preparation to preparation, the overall resulting turbidity has been very reproducible.福尔马肼悬浮液有许多特点，以确保其适合作为主要标准。它可以从试剂级的起始材料中始终如一、准确地制备。该悬浮液由不同长度和随机构型的聚合物组成，其组成的聚合物的形状和尺寸从小于0.1 μm到大于10 μm不等。尽管聚合物链长分布已被证明因制备而异，但总的浊度结果是可以很好地重现的。 5.1 Preparation of the Formazin Standards 福尔马肼标准液的制备[NOTE—All procedures described below must be performed at 20 ± 2° (see Volumetric Apparatus <31>.]• Hydrazine sulfate solution: Dissolve 1.000 g of ACS grade hydrazine sulfate (N2H4·H2SO4) in particle-free water in a 100-mL Class A volumetric flask and dilute with particle-free water to volume. Allow this solution to stand for 4–6 h.•Primary formazin standard: Dissolve 2.50 g of analytical grade hexamethylenetetramine [(CH2)6N4] in 25.0 mL of particle-free water in a 100-mL flask. Add 25.0 mL of hydrazine sulfate solution using a Class A 25-mL “to deliver” pipette and mix thoroughly. Allow the preparation to stand for 48 h at 25 ± 1° before using. The resulting suspension is stable for 2 months.•Formazin stock standard suspension 1: Using a 15-mL Class A “to deliver” pipette, transfer 15 mL of the Primary formazin standard to a 1-L volumetric flask, and dilute with particle-free water to volume and mix. The resulting suspension has a turbidity of 60 NTU.• Formazin stock standard suspension 2: Using a 50-mL Class A “to deliver” pipette, transfer 50 mL of Primary formazin standard to a 200-mL volumetric flask, and dilute with particle-free water to volume and mix. The resulting suspension has a turbidity of 1000 NTUs.• Formazin reference suspensions: Prepare by mixing in a 100-mL volumetric flask, portions of the respective Formazin stock standard suspension and particle-free water according to Table 1.[注：以下所有的程序必须在20±2°的条件下进行（参见<31 Volumetric Apparatus >）]•硫酸肼溶液：将1.000 g ACS级硫酸肼（N2H4·H2SO4）溶解在100 mL 的A类容量瓶中中，并用无颗粒水稀释至刻度。让该溶液静置4-6小时。•初级福尔马肼标准液：将2.50 g分析级六亚甲基四胺[(CH2)6N4]溶于25.0 mL无颗粒水中，装入100 mL烧瓶。使用A级25ml移液管加入25.0 mL硫酸肼溶液，并充分混合。使用前，让制剂在25±1°的温度下静置48小时。由此产生的悬浮液可稳定运行2个月。•福尔马肼储备标准悬浮液1：使用15 mL A类移液管，将15 mL福尔马肼初级标准液转移至1 L容量瓶中，并用无颗粒水稀释至刻度并混合。所得悬浮液的浊度为60 NTU。•福尔马肼储备标准悬浮液2：使用50 mL A类移液管，将50 mL福尔马肼初级标准液转移至200 mL容量瓶中，并用无颗粒水稀释至刻度并混合。所得悬浮液的浊度为1000 NTU。•福尔马肼参考悬浮液：根据表1，在100 mL容量瓶中混合各份福尔马肼储备标准悬浮液和无颗粒水，制备福尔马肼参考悬浮液。 6. QUALIFICATION OF TURBIDIMETERS AND NEPHELOMETERS 透射光式浊度仪与散射光式浊度仪的鉴定The suitability of a specific instrument for a given procedure is ensured by a stepwise life cycle evaluation for the desired application from selection to instrument retirement. The qualification comprises three components: 1) installation qualification (IQ), 2) operational qualification (OQ), and 3) performance qualification (PQ) (see Analytical Instrument Qualification <1058>).特定仪器对给定程序的适用性由从选择到仪器报废的预期应用的逐步生命周期评估来确保。鉴定包括三个部分：1）安装鉴定（IQ）、2）操作鉴定（OQ）和3）性能鉴定（PQ）（参见<1058>Analytical Instrument Qualification章节）。 The purpose of this section is to provide test methods and acceptance criteria to ensure that the instrument is suitable for its intended use (OQ), and that it will continue to function properly over extended time periods (PQ). As with any spectrometric device, a turbidimetric and nephelometric spectrometer must be qualified for both wavelength (x-axis, if not fixed) and photometric (y-axis, or signal axis) accuracy and precision, and meet the requirements for the stray light. OQ is carried out across the operational ranges required within the laboratory for both the absorbance and wavelength scales.本节的目的是提供测试方法和验收标准，以确保仪器适合其预期用途（OQ），并在延长的时间段（PQ）内继续正常工作。与任何光谱仪一样，透射光式和散射光式浊度光谱仪必须具备波长（x轴，如果不固定）和光度（y轴或信号轴）的准确度及精度，并满足杂散光的要求。OQ是在实验室内吸光度和波长标度所需的操作范围内进行的。Acceptance criteria for critical instrument parameters that establish “fitness for purpose” are verified during IQ and OQ. Specifications for particular instruments and applications can vary depending on the analytical procedure used and the desired accuracy of the final result. Instrument vendors often have samples and test parameters available as part of the IQ/OQ package.在IQ和OQ期间，验证确定“用途适用性”的关键仪器参数的验收标准。特定仪器和应用的规格可能因使用的分析程序和最终结果的预期准确度而异。仪器供应商通常将样品和测试参数作为IQ/OQ包的一部分提供。 Wherever possible in the procedures detailed as follows, primary reference standards or certified reference materials (CRMs) are to be used. Formazin is the only primary reference standard used in turbidimetry and nephelometry. All the other standards, including the CRMs, must be correlated to formazin. The CRMs should be obtained from a recognized accredited source and include independently verified traceable value assignments with associated calculated uncertainty. CRMs must be kept clean and free from dust. Recertification should be performed periodically to maintain the validity of the certification.在以下详述的程序中，应尽可能使用主要参考标准或认证参考材料（CRM）。福尔马肼是比浊法法和浊度法中唯一使用的主要参考标准。所有其他标准，包括CRM，必须与福尔马肼相关。CRM应从认可的认证来源获得，并包括独立验证的可追溯值分配及相关的计算不确定性。CRM必须保持清洁，无灰尘。应定期进行重新认证，以保持认证的有效性。 6.1 Calibration校准All of the turbidimetric and nephelometric instruments are calibrated against standards of known turbidity. The instrument must be calibrated using formazin turbidity standards prior to its first time use and at least every 3 months or as specified by the vendor. Calibration is performed using at least four formazin turbidity standards whose turbidity proportionally covers the range of interest. Many instrument manufactures provide calibration verification standards. They usually consist of sealed sample cells filled with a latex suspension or with metal oxide particles in polymer gel. These standards must be used only for checking the calibration in the time intervals between the instrument recommended calibrations.所有透射光式浊度仪和散射光式浊度仪均根据已知浊度的标准进行校准。在首次使用之前，必须使用福尔马肼浊度标准液对仪器进行校准，至少每3个月或按照供应商的规定进行一次校准。使用至少四种福尔马肼浊度标准液进行校准，其浊度按比例覆盖感兴趣的范围。许多仪器制造商提供校准验证标准。它们通常由其中充满聚合物凝胶中的金属氧化物颗粒的密封样品池或乳胶悬浮液组成。这些标准只能用于检查仪器推荐校准的时间间隔内的校准。 6.2 Stray Light杂散光Stray light (stray radiant energy) is a very significant error source, especially for measurements in the range of the lower turbidity readings. It is defined as external light that reaches the detector without being scattered from the sample. There are several sources of stray light including the inherent cell surface imperfections, reflections from within the cell that are unaccounted for, optical system parts, light sources, and, to a smaller degree, the electronics fluctuations. Although there are many design features that instrument vendors use to minimize the stray light, a complete mitigation of the stray light cannot be achieved. Unlike spectrophotometric measurements, the stray light cannot be compensated for in turbidimetry. The stray light must be measured and the values should be within the specification range set by the vendor of the particular instrument or <0.15 NTUs for the measurement in the range of 0–10 NTUs and 0.5 NTUs for the measurements in the range of 10– 1100 NTUs, whichever is smaller.杂散光(杂散光辐射能)是一个非常重要的误差源，特别是在较低的浊度读数范围内的测量。它被定义为到达探测器而不被样品散射的外部光线。杂散光有几种来源，包括电池表面固有缺陷、电池内部未被解释的反射、光学系统部件、光源，以及在较小程度上的电子波动。尽管仪器供应商使用了许多设计功能来最小化杂散光，但无法完全缓解杂散光。与分光光度测量不同，浊度法无法补偿杂散光。必须测量杂散光，其值应在特定仪器供应商设定的规格范围内，或在0-10 NTU范围内测量时小于0.15 NTU，在10-1100 NTU范围内测量时小于0.5 NTU，以较小者为准。 6.3 Range of Measuring Capability测量能力的范围The instrument must be able to measure the turbidity in the range of 0.01–1100 NTUs or from 50%–200% of the target turbidity. To demonstrate the linearity for the intended measurements range, choose at least four appropriate reference suspensions from Table 1.仪器必须能够测量0.01–1100 NTU范围内或目标浊度50%-200%范围内的浊度。为了证明预期测量范围的线性，从表1中选择至少四种合适的参考悬浮液。 6.4 Resolution 解决方案Instrument resolution must be 0.01 NTU or less for the measurements range of 0–9.99 NTUs; 0.1 NTU or less for the measurements range of 10–99.9 NTUs; and 1 NTU for the measurements above 100 NTUs.对于0-9.99 NTU的测量范围，仪器分辨率必须小于等于0.01 NTU；测量范围为10-99.9 NTU时，小于等于0.1 NTU；100 NTU以上的测量分辨率值为1 NTU。 6.5 Accuracy准确度The instrument reading accuracy must be ±10% of the reading + 0.01 NTU for the measurement range from 0–19.9 NTUs, and ±7.5% of the reading for the measurement range from 20–1100 NTUs.对于0-19.9 NTU的测量范围，仪器读数准确度必须为读数+0.01 NTU的±10%，对于20-1100 NTU的测量范围，仪器读数准确度必须为读数的±7.5%。 6.6 Performance Qualification性能鉴定The instrument PQ is accomplished periodically or as needed between the calibrations. Primary turbidity standards (formazin) or secondary calibration verification standards (latex suspensions or metal oxide particles in polymer gels contained in sealed sample cells) supplied by instrument manufacturers may be used.定期或根据需要在校准期之间完成仪器PQ。可使用仪器制造商提供的一级浊度标准（福尔马肼）或二级校准验证标准（乳胶悬浮液或密封样品池中聚合物凝胶中的金属氧化物颗粒）。 7. PROCEDURE步骤7.1 Turbidimetric Procedures 透射光比浊法测试步骤SAMPLE CELL PREPARATION 样品池准备The sample cells for sample measurements must be clean. Follow the sample cell or instrument manufacturer recommendations for cleaning the sample cells appropriately. For low turbidity measurements it is a good practice to use a single-indexed sample cell or a flow cell, which help ensure adequate precision and repeatability of the measurements. Using particle-free water, find the sample cell orientation in the sample cell holder that gives the lowest reading. For higher values of turbidity, different sample cells may be used. However, the sample cells must be matched (the difference in readings for a standard prepared at nominal sample concentration from two different sample cells must be within ±0.005 NTU or below the measurement precision requirement, whichever is lower).用于样品测量的样品室必须清洁。按照样品池或仪器制造商的建议适当清洁样品池。对于低浊度测量，最好使用一个单指数样品池或流动池，这有助于确保测量的足够精度和可重复性。使用无颗粒水，在样品池支架中找到读数最低的样品池方向。对于较高的浊度值，可使用不同的样品池。然而，样品池必须匹配（两个不同样品池在标称样品浓度下制备的标准品读数差异必须在±0.005 NTU范围内或低于测量精度要求，以较低者为准）。 SAMPLE PREPARATION样品准备Prepare the samples as prescribed in the individual monograph. Carefully mix the samples thoroughly by swirling or inverting the volumetric flask slowly several times. Avoid shaking or stirring since it may introduce bubbles. Degassing the samples helps to improve the measurements. For degassing, the samples could stand for several minutes or a vacuum could be applied, or they could be gently sonicated using an ultrasonic bath. After degassing, let the samples stand for several minutes and mix again by carefully inverting two to three times. Transfer the sample to the sample cell and take the readings.按照各专题中的规定制备样品。通过缓慢旋转或倒置容量瓶数次，仔细混合样品。避免摇晃或搅拌，因为这可能会产生气泡。对样品进行脱气有助于改进测量。对于脱气，样品可以静置几分钟，或者可以施加真空，或者可以使用超声波浴对其进行轻轻的超声波处理。脱气后，让样品静置几分钟，然后小心地反转两到三次，再次混合。将样品转移至样品池并读取读数。 USE OF FLOW CELLS流动池的使用Flow cells are mainly used for low turbidity measurements for samples with small particles. When such cells are used, the sample is introduced by carefully pouring it down the interior edge of the inlet reservoir.In practice, it is advisable to ensure that settling of the particles being measured is negligible. This is usually accomplished by including a protective colloid in the liquid-suspending medium. It is important that results be interpreted by a comparison of readings with those representing known concentrations of suspended matter, produced under precisely the same conditions.流动池主要用于小颗粒样品的低浊度测量。当使用这种样品池时，通过小心地将样品倒入进水仓的内边缘来引入样品。在实际过程中，建议确保被测颗粒的沉降可以忽略不计。这通常通过在液体悬浮介质中加入保护胶体来实现。重要的是，通过将读数与在完全相同的条件下产生的已知悬浮物浓度的读数进行比较来解释结果。 7.2 Nephelometric Procedures散射光浊度法步骤Nephelometric procedures are performed similarly to turbidimetric procedures for both direct measurements and measurements in the ratio mode as described above.散射光浊度法步骤的执行方式与透射光比浊法程序类似，适用于直接测量和上述比率模式下的测量。 RATE NEPHELOMETRIC PROCEDURES比率模式散射光浊度法步骤The overall procedure for monitoring the progress of the reaction consists of three well-defined steps: 1) record a baseline reading of the turbidity of the medium (blank); 2) record the turbidity after the first reagent (antigen) is added, which results in an increase of the turbidity until a plateau is reached; and 3) add the second reagent (antibody), which results in another turbidity increase and a second plateau followed by a final turbidity increase that continues until a third plateau is reached. The measurement zone is selected from the addition of the antibody until the third plateau, depending on the purpose of the assay and the respective component concentrations. Kinetic nephelometry and Endpoint nephelometry are two general procedures that are used for quantifying the immune complexes formed in the immunoassay methods (also known as immunonephelometry because the measured turbidity is due to immunocomplexes that are formed). For each procedure, there are several parameters that need to be optimized in each individual application. The main parameters are 1) with or without particle enhancement; 2) particle types, sizes, and respective optimum wavelength, if applicable; 3) monitoring reaction kinetic or endpoint; 4) antibody/antigen under consideration and, related to that, the optimum level of antigen loading; 5) buffers and other ionic species and respective optimal pH; 6) type and concentration of polymers used to modify the solubility of proteins; and 7) temperature and other environmental factors. Generally these parameters are optimized during the method development and the values are given in specific monograph(s) and/or chapter(s) as applicable.监测反应进程的总体程序包括三个明确定义的步骤：1）记录介质浊度的基线读数（空白）；2） 在添加一种试剂（抗原）后，记录浊度，这会导致浊度增加，直到达到一个稳定期；3）添加第二种试剂（抗体），这会导致另一个浊度增加和第二个稳定期，然后是最终浊度增加，直到达到第三个稳定器。根据分析目的和各自的组分浓度，从添加抗体到第三个稳定期中间选择测量区。动力学散射比浊法和终点散射比浊法是两种通用程序，用于量化免疫分析方法中形成的免疫复合物（也称为免疫散射比浊法，因为测得的浊度是由形成的免疫复合物引起的）。对于每一个步骤，都有几个参数需要在每个单独的应用中进行优化。主要参数为1）有无粒子增强；2） 颗粒类型、尺寸和各自的最佳波长（如适用）；3） 监测反应动力学或终点；4） 考虑中的抗体/抗原，以及与之相关的抗原负载的最佳水平；5） 缓冲液和其他离子种类以及各自的最佳pH值；6） 用于改变蛋白质溶解度的聚合物的类型和浓度；7）温度和其他环境因素。通常，这些参数在方法开发过程中进行了优化，具体的专著和/或章节（如适用）中给出了这些值。 Kinetic nephelometry: The kinetic nephelometry is advantageous compared to the endpoint nephelometry mainly because of the capability to take a sample blank reading in addition to a reagent blank reading. This procedure assesses the rate of the immunocomplex formation based on the increased intensity response of the scattered light of the chosen wavelength. The reaction kinetic may be monitored continuously or a certain number of data points may be taken, depending on the time response of the instrument used and the type of application. At times it may involve only two data points; however, proper care must be exercised because the choice of point selection can influence the overall accuracy in cases where differences in reaction kinetics exist between samples and calibrating standards. Careful consideration should be given to the appropriate choice of specificity control strategy.动力学散射比浊法：与终点散射比浊法相比，动力学散射比浊法具有优势，主要是因为除了试剂空白读数外，还能够读取样品空白读数。该程序基于所选波长的散射光的增强强度响应来评估免疫复合物的形成速率。根据所用仪器的时间响应和应用类型，可连续监测反应动力学，或采集一定数量的数据点。有时它可能只涉及两个数据点；但是在样品和校准标准之间存在反应动力学差异的情况下，选择点可能会影响整体准确度。应仔细考虑特异性控制策略的适当选择。 Endpoint nephelometry: In this method, an initial measurement is performed before adding the reagent, which represents the blank reading. A second measurement is performed after the immune complex is formed after approximately 60 min. The difference between these two measurements is proportional to the content of the component being assayed.终点散射比浊法：在该方法中，在添加试剂之前进行初始测量，这代表空白读数。大约60分钟后，在免疫复合物形成后进行第二次测量。这两次测量之间的差异与所分析成分的含量成正比。 8. VALIDATION AND VERIFICATION 验证与核查8.1 Validation 验证Validation is required when a nephelometric/turbidimetric method is intended for use as an alternative to the official procedure for testing an official article. The objective of nephelometric/turbidimetric method validation is to demonstrate that the measurement is suitable for its intended purpose, including quantitative determination of the main component in a drug substance or a drug product (Category I assays), quantitative determination of impurities or limit tests (Category II), and identification tests (Category IV). Depending on the category of the test (see Validation of Compendial Procedures <1225>, Table 2), the analytical method validation process for nephelometry/turbidimetry requires testing for accuracy, precision, specificity, detection limit (DL), quantitation limit (QL), linearity, range, and robustness. These analytical performance characteristics apply to externally standardized procedures and those that use standard additions.当散射比浊法/透射浊度法拟用作官方物品测试程序的替代方法时，需要进行验证。当散射比浊法/透射浊度法验证的目的是证明测量适用于其预期目的，包括原料药或药品中主要成分的定量测定（I类分析）、杂质的定量测定或限度试验（II类）以及鉴定试验（IV类）。根据试验的类别（参见<1225>Validation of Compendial Procedures，表2），透射浊度法/散射比浊法的分析方法验证过程需要对准确度、精密度、特异性、检测限（DL）、定量限（QL）、线性、范围和稳健性进行试验。这些分析性能特征适用于外部标准化程序和那些使用标准添加的程序。 Validation of Compendial Procedures <1225> provides definitions and general guidance on analytical procedures validation without indicating specific validation criteria for each characteristic. The intention of the following sections is to provide the user with specific validation criteria that represent the minimum expectations for this technology. For each particular application, tighter criteria may be needed in order to demonstrate suitability for the intended use.<1225>Validation of Compendial Procedures章节提供了分析程序验证的定义和一般指南，但没有说明每个特征的具体验证标准。以下各节的目的是向用户提供具体的验证标准，这些标准代表了对该技术的最低期望。对于每个特定应用，可能需要更严格的标准，以证明其适用于预期用途。 ACCURACY 准确度For Category I, II, and III procedures, accuracy can be determined by conducting recovery studies with the appropriate matrix spiked with known concentrations of the analyte. Analysts can also compare the assay results obtained using the nephelometric/ turbidimetric procedure under validation to those from an established analytical procedure.Validation criteria: 98.0%–102.0% mean recovery for the drug substances, 95.0%–105.0% mean recovery for the drug product assay, and 80.0%–120.0% mean recovery for the impurity analysis. These criteria are met throughout the specified range. 对于I类、II类和III类程序，可通过使用加入已知分析物浓度的适当基质进行回收研究来确定准确度。分析员还可以将使用验证中的散射光浊度法/透射光比浊法程序获得的分析结果与已建立的分析程序获得的结果进行比较。验证标准：原料药的平均回收率为98.0%–102.0%，药品分析的平均回收率为95.0%–105.0%，杂质分析的平均回收率为80.0%–120.0%。这些标准在整个规定范围内都得到满足。 PRECISION精度Repeatability: The repeatability of the analytical procedure is assessed by measuring the concentrations of six independently prepared sample solutions at 100% of the assay test concentration. Alternatively, it can be assessed by measuring the concentrations of three replicates of three separate sample solutions at different concentrations. The three concentrations should be close enough so that the repeatability is constant across the concentration range. If this is done, the repeatability at the three concentrations is pooled for comparison to the acceptance criteria.Validation criteria: The relative standard deviation is NMT 1.0% for the drug substance, NMT 2.0% for the drug product assay, and NMT 20.0% for the impurity analysis.重复性：通过测量六种独立制备的样品溶液在100%分析试验浓度下的浓度来评估分析程序的重复性。或者，可以通过测量三种不同浓度的单独样品溶液的三个重复的浓度来评估。三种浓度应足够接近，以便在整个浓度范围内重复性保持恒定。如果这样做，将三种浓度下的重复性汇总，以与验收标准进行比较。验证标准：原料药的相对标准偏差为NMT 1.0%，药品分析的相对标准偏差为NMT 2.0%，杂质分析的相对标准偏差为NMT 20.0%。 Intermediate precision: The effect of random events on the analytical precision of the method must be established. Typical variables include performing the analysis on different days, using different instrumentation, and/or having the method performed by two or more analysts. At a minimum, any combination of at least two of these factors totaling six experiments will provide an estimation of intermediate precision.Validation criteria: The relative standard deviation is NMT 1.5% for the drug substance, NMT 3.0% for the drug product assay, and NMT 25.0% for the impurity analysis.中间精度：必须确定随机事件对方法分析精度的影响。典型的变量包括在不同的日期使用不同的仪器进行分析，和/或由两名或两名以上的分析员进行分析。至少，这些因素中的至少两个的组合，总共6个实验，将提供中等精度的评估。验证标准：原料药的相对标准偏差为NMT 1.5%，药品分析的相对标准偏差为NMT 3.0%，杂质分析的相对标准偏差为NMT 25.0%。 SPECIFICITY 特异性In nephelometric/turbidimetric measurements, specificity is demonstrated by the lack of interference from other components present in the matrix (other components of the matrix produce a true solution).在散射光浊度法/透射光比浊法的浊度测量中，特异性通过基质中其他成分的干扰（基质的其他成分产生真实溶液）的缺乏来证明。 DETECTION LIMIT 检测限The DL can be estimated by calculating the concentration of a solution that would give the signal-to-noise ratio of ≥3.3. The estimated DL must be confirmed by analyzing samples at the calculated concentration.可以通过计算溶液的浓度来估计检测限DL，该浓度将给出信号的信噪比≥3.3. 必须通过分析计算浓度下的样品来确认估计的DL。 QUANTITATION LIMIT 定量限The QL can be estimated by calculating the concentration of a solution that would give the signal-to-noise ratio of ≥10.0. The estimated QL must be confirmed by analyzing samples at the calculated concentration. Measurement of a test solution prepared from a representative sample matrix spiked at the required QL concentration must be performed to confirm sufficient sensitivity and adequate precision. The observed signal-to-noise ratio at the required QL should be >10.Validation criteria: For the estimated limit of quantitation to be considered valid, the measured concentration must be accurate and precise at a level ≤50% of the specification.定量限QL可以通过计算溶液的浓度来估算，该浓度将给出信号的信噪比≥10.0. 必须通过分析计算浓度下的样品来确认估算的QL。必须对以所需QL浓度添加的代表性样品基质制备的试液进行测量，以确认其具有足够的灵敏度和精度。在所需QL下观察到的信噪比应大于10。验证标准:估计的定量限被认为是有效的，测量的浓度必须是准确的，并且在≤50%的规格水平上是精确的。 LINEARITY线性A linear relationship between the analyte concentration and measured turbidity response must be demonstrated by preparation of at least four standard solutions at concentrations encompassing the anticipated concentration of the test solution. The standard curve is then evaluated using appropriate statistical methods such as a least-squares regression. Deviation from linearity results from instrumental or sample factors, or both, can be reduced to acceptable levels by reducing or increasing the analyte concentration, thereby respectively decreasing or increasing the turbidity readings to within the nephelometer/turbidimeter instrument linearity range.Validation criteria: The correlation coefficient (R) must be NLT 0.995 for Category I assays and NLT 0.99 for Category II quantitative tests.分析物浓度和测得的浊度响应之间的线性关系必须通过制备至少四种标准溶液来证明，其浓度包括试验溶液的预期浓度。然后使用适当的统计方法（如最小二乘回归）评估标准曲线。通过降低或增加分析物浓度，可将仪器或样品因素或两者的线性偏差降低至可接受水平，从而分别将浊度读数降低或增加至透射光法浊度计/散射光浊度计仪器线性范围内。验证标准：对于I类分析，相关系数（R）必须为NLT 0.995，对于II类定量测试，相关系数（R）必须为NLT 0.99。 RANGE 范围The operational range of an analytical instrument (and the analytical procedure as a whole) is the interval between the upper and lower concentrations (amounts) of analyte in the sample (including these concentrations) for which it has been demonstrated that the instrumental response function has a suitable level of precision, accuracy, and linearity.Validation criteria: For Category I tests, the validation range for 100.0% centered acceptance criteria is 80.0%–120.0%. For non-centered acceptance criteria, the validation range is 10.0% below the lower limit to 10.0% above the upper limit. For Category II tests, the validation range covers 50.0%–120.0% of the acceptance criteria.分析仪器（以及整个分析程序）的操作范围是样品中分析物的上下浓度（数量）（包括这些浓度）之间的间隔，已证明仪器响应函数具有适当的精度、准确度和线性水平。验证标准：对于I类试验，100.0%中心验收标准的验证范围为80.0%–120.0%。对于非中心验收标准，验证范围为下限以下10.0%到上限以上10.0%。对于II类试验，验证范围涵盖验收标准的50.0%–120.0%。 ROBUSTNESS稳健性The reliability of an analytical measurement is demonstrated by deliberate changes to experimental parameters. For nephelometry/turbidimetry this can include, for example, measuring the stability of the analyte under specified storage conditions, varying pH, and adding possible interfering species. Robustness is determined concurrently using a suitable design for the experimental procedure.分析测量的可靠性通过有意改变实验参数来证明。 对于散射光浊度法/透射光比浊法，这可以包括：测量分析物在特定储存条件、变化的 pH 值和添加可能的干扰物质下的稳定性。使用适合实验程序的设计，同时确保稳健性。 8.2 Verification核查Current U.S. Good Manufacturing Practices regulations [21 CFR 211.194(a)(2)] indicate that users of analytical procedures described in the U.S. Pharmacopeia and National Formulary are not required to validate these procedures if provided in a monograph. Instead, they simply must verify their suitability under actual conditions of use.现行的《美国生产规范条例》[21 CFR 211.194（a）（2）]表明，如果专论中提供了这些程序，则美国药典和国家处方集中描述的分析程序的用户无需验证这些程序。相反，他们只需验证其在实际使用条件下的适用性。 The objective of nephelometric/turbidimetric procedure verification is to demonstrate the suitability of a test procedure under actual conditions of use. Performance characteristics that verify the suitability of a nephelometric/turbidimetric procedure are similar to those required for any analytical procedure. A discussion of the applicable general principles is found in Verification of Compendial Procedures <1226>. Verification is usually performed using a reference material and a well-defined matrix. Verification of compendial nephelometric/turbidimetric procedures includes, at minimum, the execution of the validation parameters for specificity, accuracy, precision, and QL, when appropriate, as indicated in 8.1 Validation.散射光浊度法/透射光比浊法程序验证的目的是证明测试程序在实际使用条件下的适用性。验证散射光浊度法/透射光比浊法程序适用性的性能特征与任何分析程序所需的性能特征相似。适用的一般原则的讨论见<1226>Verification of Compendial Procedure 章节。通常使用参考材料和明确定义的基质进行验证。 药典散射光浊度法/透射光比浊法程序的验证至少包括对特异性、准确度、精密度和 QL 的验证参数的执行（如 8.1 验证中所述）。 欧洲药典EP10.02.2.1. CLARITY AND DEGREE OF OPALESCENCE OF LIQUIDS 液体的澄清度和乳光度Opalescence is the effect of light being absorbed or scattered by submicroscopic particles or optical density inhomogeneities. The absence of any particles or inhomogeneities in a solution results in a clear solution.光被亚微观粒子吸收或散射、或光密度不均匀的产生的效果即为乳光。溶液中不存在任何粒子或不均匀性，就会得到清澈的溶液。 A liquid is considered clear if its clarity is the same as that of water R or of the solvent used, or if its opalescence is not more pronounced than that of reference suspension I (see Table 2.2.1.-1), when examined under the conditions described below.在下述条件下检查时，如果液体的透明度与水或所用溶剂的透明度相同，或者其乳光不比参考悬浮液I（见表2.2.1.-1）的乳光更明显，则认为液体是透明的。 Requirements in monographs are expressed in terms of the visual method by comparing with the defined reference suspensions (see Table 2.2.1.-1). However, instrumental methods may also be used for determining compliance with monograph requirements once the suitability of the instrument has been established as described below and calibration with reference suspensions I-IV and with water R or the solvent used has been performed.通过与规定的参考悬浮液进行比较（见表2.2.1.-1），以目视法表达专著中的要求。然而，一旦仪器的适用性如下所述建立，仪器方法也可用于确定是否符合专论要求，并使用参考悬浮液I-IV和水或所用溶剂进行校准。VISUAL METHOD目视法Using identical test-tubes of colourless, transparent, neutral glass with a flat base and an internal diameter of 15-25 mm, compare the liquid to be examined with a reference suspension freshly prepared as described below. Ensure that the depths of the layers in the 2 test-tubes are the same (about 40 mm).使用相同的无色透明中性玻璃试管，底座平坦，内径为15-25 mm，将待检液体与下述新制备的参考悬浮液进行比较。确保两个试管中各层的深度相同（约40 mm）。 Compare the liquids in diffused daylight 5 min after preparation of the reference suspension, viewing vertically against a black background.制备参考悬浮液5分钟后，在漫射日光下比较液体，在黑色背景下垂直观察。 System suitability. The diffusion of light must be such that reference suspension I can readily be distinguished from water R, and that reference suspension II can readily be distinguished from reference suspension I (see Table 2.2.1.-1).系统适用性。光的扩散必须确保参考悬浮液I可以很容易地与水区分开，并且参考悬浮液II可以很容易地与参考悬浮液I区分开（见表2.2.1.-1）。INSTRUMENTAL METHOD 仪器法The instrumental assessment of clarity and opalescence provides a more discriminatory test that does not depend on the visual acuity of the analyst. Numerical results are more useful for process control and quality monitoring, especially in stability studies. For example, previous numerical data on stability can be extrapolated to determine whether a given batch of a preparation will exceed shelf-life limits prior to the expiry date.仪器法评估给透明度和乳光度的提供了一种更具辨别力的测试，它不依赖于分析人员的视力。 数值结果对于过程控制和质量监控更有用，尤其是在稳定性研究中。 例如，可以从以前关于稳定性的数字数据外推，来确定给定批次的制剂是否会在有效期之前超过保质期限制。 TURBIDIMETRY AND NEPHELOMETRY比浊法和浊度法When a suspension is viewed at right angles to the direction of the incident light, the system appears opalescent due to the scattering of light by the particles of the suspension (Tyndall effect). A certain portion of the light beam entering a turbid liquid is transmitted, another portion is absorbed and the remaining portion is scattered by the suspended particles. The light-scattering effect of suspended particles can be measured either indirectly by observation of the transmitted light (turbidimetry) or directly by measuring the scattered light (nephelometry). Turbidimetry and nephelometry are more reliable in low turbidity ranges, where there is a linear relationship between turbidity values and detector signals. As the degree of turbidity increases, not all the particles are exposed to the incident light and the scattered or the transmitted radiation of other particles is hindered on its way to the detector.当以与入射光方向成直角的角度观察悬浮液时，由于悬浮液颗粒对光的散射（丁达尔效应），系统呈现乳白色。进入混浊液体的光束的一部分被透射，另一部分被吸收，其余部分被悬浮颗粒散射。悬浮颗粒的光散射效应可以通过观察透射光（比浊法）间接测量，也可以通过测量散射光（浊度法）直接测量。比浊法和浊度法在低浊度范围内更可靠，浊度值和检测器信号之间存在线性关系。随着浊度的增加，并非所有粒子都暴露在入射光下，其他粒子的散射或透射辐射在到达探测器的过程中会受到阻碍。 For quantitative measurements, the construction of calibration curves is essential. Linearity must be based on at least 4 levels of concentrations. Reference suspensions must show a sufficiently stable degree of turbidity and must be produced under well-defined conditions.对于定量测量，校准曲线的构建至关重要。线性必须基于至少4个浓度水平。参考悬浮液必须显示足够稳定的浊度，并且必须在明确的条件下产生。 MEASUREMENTS IN RATIO MODE比率模式下的测量The determination of opalescence of coloured liquids is done using instruments with ratio mode, since colour provides a negative interference, attenuating both incident and scattered light and lowering the turbidity value. The effect is so great, even for moderately coloured samples, that conventional nephelometers cannot be used.由于颜色会产生负干扰，衰减入射光和散射光，降低浊度值，因此使用具有比率模式的仪器测定有色液体的乳光。 这种影响是如此之大，即使是中等颜色的样品，以至于不能使用传统的浊度计。 In turbidimetry or nephelometry with ratio mode, the ratio of the transmission measurement to the 90° scattered light measurement is determined. This procedure compensates for the light that is diminished by the colour of the sample. Instruments with ratio mode use as light source a tungsten lamp with spectral sensitivity at about 550 nm operating at a filament colour temperature of 2700 K. Other suitable light sources may also be used. Silicon photodiodes and photomultipliers are commonly used as detectors and record changes in light scattered or transmitted by the sample. The light scattered at 90 ± 2.5° is measured by the primary detector. Other detectors measure back and forward scatter (reflected light) as well as transmitted light. The results are obtained by calculating the ratio of the 90° scattered light measured to the sum of the components of forward scattered and transmitted light values.在比浊法或浊度法中，通过比率模式，确定透射测量与90°散射光测量的比率。该程序补偿因样品颜色而减弱的光线。具有比率模式的仪器使用光谱灵敏度约为550 nm的钨灯作为光源，在2700 K的灯丝色温下工作。也可以使用其他合适的光源。硅光电二极管和光电倍增管常用作探测器，记录样品散射或透射光的变化。由主探测器测量90±2.5°处的散射光。其他探测器测量前后散射（反射光）以及透射光。通过计算测得的90°散射光与前向散射光和透射光值分量之和的比值，可以获得结果。 The instruments used are calibrated against standards of known turbidity and are capable of automatic measurement of turbidity. The test results are obtained directly from the instrument and compared to the specifications in the individual monograph.使用的仪器根据已知浊度标准进行校准，并能够自动测量浊度。测试结果直接从仪器中获得，并与各专著中的规范进行比较。 Alternatively, the influence of the colour of the sample may also be eliminated by using an infrared light-emitting diode (IR LED) having an emission maximum at 860 nm with a 60 nm spectral bandwidth as the light source of the instrument.或者，也可以通过使用最大发射波长为860nm、光谱带宽为60nm的红外发光二极管（IR LED）作为仪器光源来消除样品颜色的影响。 INSTRUMENT REQUIREMENTS仪器要求Instruments complying with the following characteristics and verified using reference suspensions as described below may be used instead of visual examination for determination of compliance with monograph requirements.可使用符合以下特征并使用下述参考悬浮液验证的仪器代替目视检查，以确定是否符合专论要求。 – Measuring unit: NTU (nephelometric turbidity units). NTU is based on the turbidity of a primary standard of formazin. FTU (formazin turbidity units) or FNU (formazin nephelometric units) are also used, and are equivalent to NTU in regions of low turbidity (up to 40 NTU). These units are used in all 3 instrumental methods (nephelometry, turbidimetry and in ratio mode).– Measuring range: 0.01-1100 NTU.– Resolution: 0.01 NTU within the range 0-9.99 NTU; 0.1 NTU within the range 10.0-99.9 NTU; and 1 NTU for the range > 100 NTU.– Accuracy: ± (10 per cent of reading + 0.01 NTU) with in the range 0-20 NTU; ± 7.5 per cent within the range 20-1100 NTU.– Repeatability: ± 0.05 NTU within the range0-20 NTU; ± 2 per cent of the reading within the range 20-1100 NTU.测量单位：NTU（浊度测量单位）。NTU是基于福尔马肼一级标准品的浊度。也可使用FTU（福尔马肼浊度单位）或FNU（福尔马肼浊度单位），相当于NTU的在低浊度区域（最高40 NTU）。这些单位适用于所有3种仪器方法（比浊法、浊度法和比率模式）。–测量范围：0.01-1100 NTU–分辨率：0-9.99 NTU范围内为0.01 NTU；10.0-99.9 NTU范围内为0.1 NTU；对于大于100 NTU的范围，则为1 NTU–准确度：范围在0-20 NTU之间，读数准确度偏差为±（读数的10%+0.01 NTU）；范围在20-1100 NTU时，读数准确偏差为±7.5%。–重复性：在0-20 NTU范围内重复性为±0.05 NTU；在20-1100 NTU范围内读数重复性为±2%。 Instruments with measuring range or resolution, accuracy and repeatability capabilities other than those mentioned above may be used provided they are sufficiently validated and are capable for the intended use.测量范围或分辨率、精度和重复性能力不同于上述测量范围或分辨率、精度和重复性能力的仪器经过有效验证，也能够应用于预期用途。 CONTROL OF INSTRUMENT PERFORMANCE仪器性能的控制– Calibration: performed with at least 4 reference suspensions of formazin covering the measuring range of interest. Reference suspensions described in this chapter or suitable reference standards calibrated against the primary reference suspensions may be used.–校准：使用至少4种福尔马肼参考悬浮液进行校准，覆盖感兴趣的测量范围。可使用本章所述的参考悬浮液或根据主要参考悬浮液校准的适当参考标准。 – Stray light: < 0.15 NTU within the range 0-10 NTU; < 0.5 NTU within the range 10-1100 NTU. Stray light is defined as that light that reaches the nephelometric detector without being a result of scatter from the sample. Stray light is always a positive interference and is a significant source of error in low-range turbidity measurements. Sources of stray light include: imperfections in and scratches on sample cells, internal reflections of the optical system, contamination of the optics or sample cell chamber with dust, and electronic noise. Instrument design can also affect stray light. The influence of stray light becomes negligible in ratio mode measurements.杂散光：在0-10NTU范围内<0.15 NTU；在10-1100 NTU范围内<0.5 NTU。杂散光是指到达浊度检测器的光，而不是样品散射的结果。杂散光总是一种正干扰，是低范围浊度测量中的一个重要误差源。杂散光的来源包括：样品池中的缺陷和划痕、光学系统的内部反射、光学元件或样品池被灰尘污染，以及电子噪声。仪器设计也会影响杂散光。在比率模式测量中，杂散光的影响可以忽略不计。 The test methodology for the specific substance/product to be analysed must also be verified to demonstrate its analytical capability. The instrument and methodology shall be consistent with the attributes of the substance to be examined.还必须验证待分析特定物质/产品的试验方法，以证明其分析能力。仪器和方法应与待检物质的属性一致。 Measurements of standards and samples should be carried out under the same temperature conditions, preferably between 20 °C and 25 °C.标准品和样品的测量应在相同的温度条件下进行，最好在20 °C和25 °C之间。 REFERENCE SUSPENSIONS 参考悬浮液Formazin has several desirable characteristics that make it an excellent turbidity standard. It can be reproducibly prepared from assayed raw materials. The physical characteristics make it a desirable light-scatter calibration standard. The formazin polymer consists of chains of different lengths, which fold into random configurations. This results in a wide variety of particle shapes and sizes, which allows the analysis of different particle sizes and shapes that are found in real samples. Stabilised formazin suspensions that can be used to prepare stable, diluted turbidity standards are commercially available and may be used after comparison with the standards prepared as described.福尔马肼有几个理想的特性，使其成为一个优秀的浊度液标准。它可以从经过分析的原材料中重复制备。其物理特性使其成为理想的光散射校准标准。福尔马肼聚合物由不同长度的链组成，这些链折叠成随机构型。这会产生各种各样形状和尺寸的颗粒，从而可以分析真实样品中发现的不同颗粒大小和形状。可用于制备稳定稀释浊度标准品的稳定福尔马肼悬浮液是可商购的，并可在与所述制备的标准品进行比较后使用。 All steps of the preparation of reference suspensions as described below are carried out at 25 ± 3 °C.下述参考悬浮液制备的所有步骤均在25±3°C下进行。 Hydrazine sulfate solution. Dissolve 1.0 g of hydrazine sulfate R in water R and dilute to 100.0 mL with the same solvent. Allow to stand for 4-6 h.硫酸肼溶液。 将 1.0 g 硫酸肼溶解在水中，并用相同的溶剂稀释至100.0 mL。 静置 4-6 小时。 Primary opalescent suspension (formazin suspension). In a 100 mL ground-glass-stoppered flask, dissolve 2.5 g of hexamethylenetetramine R in 25.0 mL of water R. Add 25.0 mL of the hydrazine sulfate solution. Mix and allow to stand for 24 h. This suspension is stable for 2 months, provided it is stored in a glass container free from surface defects. The suspension must not adhere to the glass and must be mixed thoroughly before use.初级乳白色悬浮液（福尔马肼悬浮液）。在100 mL磨砂玻璃塞烧瓶中，将2.5g六亚甲基四胺溶解在25.0 mL水中。添加25.0 mL硫酸肼溶液。混合并静置24小时。如果该悬浮液储存在无表面缺陷的玻璃容器中，则可稳定2个月。悬浮液不得粘附在玻璃上，使用前必须彻底混合。 Standard of opalescence. Dilute 15.0 mL of the primary opalescent suspension to 1000.0 mL with water R. This suspension is freshly prepared and may be stored for up to 24 h.乳白色的标准浊度液。用水将15.0 mL初级乳白色悬浮液稀释至1000.0 mL。该悬浮液是新制备的，可储存24小时。 Reference suspensions. Prepare the reference suspensions according to Table 2.2.1.-1. Mix and shake before use.参考悬浮液。根据表2.2.1-1制备参考悬浮液。使用前混合并摇匀。Measurements of reference suspensions I-IV in ratio mode show a linear relationship between the concentrations and measured NTU values (see Table 2.2.1.-2). 在比率模式下，参考悬浮液I-IV的测量结果显示，浓度与测量的NTU值之间存在线性关系（见表2.2.1.-2）。日本药典17版2.61 Turbidity Measurement 浊度测量Turbidity measurement is used to determine the turbidity (degree of opalescence) for the decision whether the article to be examined complies with the clarity requirement stated in the Purity.As a rule, the visual method is specified for the requirement in individual monograph.浊度测量用于确定浊度（乳光度），以决定待检查的物品是否符合纯度中规定的透明度要求。作为一项规则，目视法是针对个别专论中的要求说明的。 1. Visual method目视法This is used to determine the degree of opalescence with white (or faintly-colored) fine particles. So the degree of opalescence of a colored sample is liable to be determined lower that it is difficult to compare the degree correctly without using similarly colored reference suspension.这是用来确定乳白色(或淡色)细颗粒的乳光程度。因此，有色样品的乳光度容易被测定得较低，因此，如果不使用类似颜色的参考悬浮液，就很难正确地比较其乳光度。 1.1. Reference suspensions参考悬浮液Pipet 5 mL, 10 mL, 30 mL and 50 mL of formazin opalescence standard solution, dilute them separately to exactly 100 mL with water, and use these solutions so obtained as Reference suspensions I, II, III and IV, respectively. Shake before use. Degrees of opalescence of Reference suspensions I, II, III and IV are equivalent to 3 NTU, 6 NTU, 18 NTU and 30 NTU, respectively.用移液管分别吸取5 mL、10 mL、30 mL、50 mL福尔马肼标准液，用水分别稀释至100 mL，分别作为参比悬液I、II、III、IV。在使用前摇晃。参考悬浮液I、II、III和IV的乳光度分别相当于3 NTU、6 NTU、18 NTU和30 NTU。 1.2. Procedure步骤Place sufficient of the test solution, water or the solvent to prepare the test solution and, where necessary, newly prepared Reference suspensions in separate flat-bottomed test tubes, 15 – 25 mm in inside diameter and of colorless and transparent, to a depth of 40 mm, and compare the contents of the tubes against a black background by viewing in diffused light down the vertical axes of the tubes. The diffused light must be such that Reference suspension I can be readily distinguished from water, and that Reference suspension II can readily be distinguished from Reference suspension I.取足够的待测溶液、水或溶剂，以准备测试溶液，必要时，将新制备的参考悬浮液置于独立的平底试管中，试管内径15 - 25mm，无色透明，深度40 mm。然后在一个黑色的背景下通过漫射光下垂直于管轴进行观察，比较管内的内容。漫射光必须能使参考悬浮体I容易与水区分开来，参考悬浮体II容易与参考悬浮体I区分开来。 In this test Reference suspensions are used when the clarity of the test solution is obscurely and it is not easy to determine that its degree of opalescence is similar or not similar to water or to the solvent used to prepare the test solution.在此测试中，当测试溶液的透明度模糊不清，并且不容易确定其乳光度与水或与用于制备测试溶液的溶剂是否相似时，使用参考悬浮液。 1.3. Interpretation注释A liquid is considered “clear” when its clarity is the same as that of water or of the solvent used to prepare the liquid or its turbidity is not more pronounced than that of Reference suspension I. If the turbidity of the liquid is more than that of Reference suspension I, consider as follows: When the turbidity is more than that of Reference suspension I but not more than that of Reference suspension II, express “it is not more than Reference suspension II”. In the same way, when the turbidity is more than that of Reference suspension II but not more than that of Reference suspension III, express “it is not more than Reference suspension III”, and when the turbidity is more than that of Reference suspension III but not more than that of Reference suspension IV, express “it is not more than Reference suspension IV”. When the turbidity is more than that of Reference suspension IV, express “it is more than Reference suspension IV”.当液体的澄清度与水或与用于制备液体的溶剂的澄清度相同或其浊度不比参比悬浮液 I 更明显时，该液体被视为“澄清”。如果液体的浊度大于参考悬浮液I，考虑如下：当浊度大于参考悬浮液I但不超过参考悬浮液II时，表示“不超过参考悬浮液II”。 同理，当浊度大于参比悬浊液Ⅱ但不大于参比悬浊液Ⅲ时，表示“不大于参比悬浊液Ⅲ”，当浊度大于参比悬浊液Ⅲ时 但不超过参考悬浮液IV，表示“不超过参考悬浮液IV”。 当浊度大于参考悬浮液IV时，表示“大于参考悬浮液IV”。 1.4. Reagent solutions试剂溶液Formazin opalescence standard solution: To exactly 3 mL of formazin stock suspension add water to make exactly 200 mL. Use within 24 hours after preparation. Shake thoroughly before use. Degrees of opalescence of this standard solution is equivalent to 60 NTU.福尔马津乳光标准溶液：准确地取 3 mL福尔马肼储备悬浮液，加水至 200 mL。 配制后24小时内使用。使用前彻底摇匀。此标准溶液的乳光度相当于 60 NTU。 2. Photoelectric photometry光电光度法The turbidity can also be estimated by instrumental measurement of the light absorbed or scattered on account of submicroscopic optical density inhomogeneities of opalescent solutions and suspensions. The photoelectric photometry is able to provide more objective determination than the visual method. Though they can determine the turbidity by measuring the scattered or transmitted light, the measuring system and light source must be specified in individual test method, and for the comparison of observed data, the same measuring system and light source should be used.由于乳光溶液和悬浮液的亚显微光密度不均匀性，还可以通过仪器测量吸收或散射的光来估计浊度。光电光度法比目测法能够提供更客观的测定。 虽然他们可以通过测量散射光或透射光来确定浊度，但必须在单独的测试方法中标明测量系统和光源，并且为了比较观察数据，应使用相同的测量系统和光源。 In each case, the linear relationship between turbidity and concentration must be demonstrated by constructing a calibration curve using at least 4 concentrations. For colored samples, the turbidity value is liable to be estimated lower because of attenuating both incident and scattered lights due to the absorption by the color, and the transmission-dispersion method is principally used.在每种情况下，浊度和浓度之间的线性关系必须通过使用至少4种浓度构建校准曲线来证明。对于有颜色的样品，由于颜色的吸收，入射光和散射光都被衰减，浊度值容易被估计得较低，主要采用透射-色散法。 2.1. Turbidimetry透射光比浊法When a light passes through a turbid liquid the transmitted light is decreased by scattering with the particles dispersed in the liquid. A linear relationship is observed between turbidity and concentration when the particles with a constant size are uniformly dispersed, the size is small and the suspension is not higher concentration. The turbidity can be measured by Ultraviolet-visual Spectrophotometry <2.24> using spectrophotometer or photoelectric photometer. The turbidity of the sample in higher concentration can also be measured, however, it is susceptible to the color of the sample, and the measurement is usually performed at around 660 nm to avoid possible disturbance occurred from the absorption by the color.当光通过混浊液体时，透射光通过分散在液体中的颗粒散射而减少。当粒径恒定的颗粒分散均匀、粒径较小且悬浮液浓度不高时，浊度与浓度呈线性关系。浊度可以通过紫外分光光度法<2.24>使用分光光度计或光电光度计进行测量。较高浓度的样品的浊度也可以测量，但它易受样品颜色的影响，通常在660 nm左右进行测量，以避免颜色吸收可能产生的干扰。 2.2. Nephelometry散射光浊度法When a suspension is viewed at right angles to the direction of the incident light, it appears opalescent due to the refraction of light from the particles of the suspension (Tyndall effect). A certain portion of the light entering a turbid liquid is transmitted, another portion is absorbed and the remaining portion is scattered by the suspended particles. The scattered light measuring method shows the linear relationship between the nephelometric turbidity units (NTU) values and relative detector signals in a low turbidity range. As the degree of turbidity increases, not all the particles are exposed to the incident light and the scattered radiation of other particles is hindered on its way to the detector.当悬浮物与入射光方向成直角时，由于悬浮物粒子的光线折射(丁达尔效应)，悬浮物呈现乳白色。进入混浊液体的光，一部分被透射，一部分被吸收，剩下的部分被悬浮的粒子散射。散射光测量方法显示了低浊度范围内散射浊度单位(NTU)值与相对检测器信号之间的线性关系。随着浊度的增加，并不是所有的粒子都暴露在入射光下，其他粒子的散射辐射在到达探测器的过程中会受到阻碍。 2.3. Ratio Turbidimetry比率浊度法This method measures both scattered and transmitted light values at the same time, and the turbidity is determined from the ratio of the scattered light value to the transmitted light value. This procedure compensates for the light that is diminished by the color of the sample and eliminates the influence of the color. When the measurement is performed by using an integrating sphere, it is particularly called the integrating sphere method, which measures the total transmitted light value as well as the scattered light value occurred with the suspended particles, and the turbidity can be determined from the ratio of them.该方法同时测量散射光值和透射光值，浊度由散射光值与透射光值的比值确定。此程序可补偿因样品颜色而减弱的光线，并消除颜色的影响。当用积分球进行测量时，特别称为积分球法，它测量悬浮粒子的总透射光值和散射光值，由它们的比值可以确定浊度。 2.4. Application of photoelectric photometry for monograph requirements光电光度法在专著要求中的应用The turbidity of the test solution, determined by the photoelectric photometry, can be used as an indicating standard for the conformity to the clarity requirements by converting into NTU by using turbidity known reference solutions such as Reference suspensions I – IV, if needed, and water or the solvent used. In an automatically compensable apparatus being calibrated with turbidity known reference solutions, the measuring result is given in NTU and it can be compared directly with required specified value.由光电光度法测定的测试溶液的浊度，可以作为指示标准，通过使用浊度已知的参考溶液，如参考悬浮液 I-IV，如果需要，水和使用的溶剂液也可以，将其以NTU为单位的数据转出。在使用浊度已知参考溶液校准的自动补偿装置中，测量结果以 NTU 为单位给出，并且可以直接与所需的规定值进行比较。 NTU is often used as the unit in the turbidity determinations. It is the unit used in the case when the turbidity is estimated by the instrument which measures the 90 ± 30°scattered light against the incident light intensity, using tungsten lamp, and in the case the estimation is performed by the instrument which measures the 90 ± 2.5°scattered light against the incident light intensity using 860 nm infrared light, FNU is used as the unit. FNU is equivalent to NTU at a range of smaller measurements (less than 40 NTU). For the unit of formazin concentration, FTU is also used, which is defined as a suspension of 1 mg formazin in 1L of purified water is 1 FTU.在浊度测定中经常使用NTU作为单位。它是测量用90±30°散射光对入射光强度的得到浊度信息时使用的单位，使用钨灯，采用860 nm红外光测量90±2.5°的散射光对入射光强度，此时以FNU为单元。在较小的测量范围内(小于40 NTU)，FNU相当于NTU。福尔马肼的浓度单位也用FTU，即1L纯净水中1 mg 福尔马肼的悬浮液为1 FTU。 Formazin stock suspension. To 25 mL of hexamethylenetetramine TS add 25 mL of hydrazinium sulfate TS, mix, and use after allowing to stand at room temperature for 24 hours. Store in a glass container free from surface defects. Use within 2 months. Shake thoroughly before use. The turbidity of this suspension is equivalent to 4000 NTU.福尔马肼贮备悬浮液：向 25 mL六亚甲基四胺中加入25 mL硫酸肼，混匀，室温静置 24 小时后使用。 储存在没有表面缺陷的玻璃容器中。2个月内使用。使用前彻底摇匀。这种悬浮液的浊度相当于4000 NTU。 Formazin opalescence standard solution. To 15 mL of formazin stock suspension add water to make 1000 mL. Use within 24 hours after preparation. Shake thoroughly before use.福尔马肼标准液：向 15 mL 的福尔马肼贮备悬浮液中加水至 1000 mL。配制后24小时内使用。使用前彻底摇匀。 解决方案：上海胤煌科技针对药剂的澄清度检查推出了以下产品，符合各国药典的溶液澄清度检查规范。1、澄清度检查专用伞棚灯HN-100A型和HN-200A型澄清度检查专用伞棚灯符合各国药典中目视法检测溶液澄清度的仪器要求，具有光林带型光源，能有效减少目视过程中光对眼睛的刺激，其照度可达5000 Lux。其中HN-200A型专用伞棚灯增加了RGB三色光源，可以对有色样品进行澄清度检测。 2、YH-CLS-1201澄清度检查分析仪此仪器采用全彩液晶触摸屏进行操作控制，可以直接检测注射用原料药和注射剂的澄清度，并具备四级权限管理和审计追踪功能，完全满足GMP的数据完整性要求，是液体一致性评价的有效仪器。