USP38-NF33_C71_无菌检查法-中英对照

USP38 NF33

<71> STERILITY TESTS

<71>无菌检查法

◆Portions of this general chapter have been harmonized with the corresponding texts of the European Pharmacopeia and/or the Japanese Pharmacopeia. Those portions that are not harmonized are marked with symbols (◆◆) to specify this fact.◆

◆本检查法已与《欧洲药典》和《日本药局方》对应部分进行了协调，不一致的

部分用符号()标注。◆

These Pharmacopeial procedures are not by themselves designed to ensure that a batch of product is sterile or has been sterilized. This is accomplished primarily by validation of the sterilization process or of the aseptic processing procedures.

按药典规定的无菌检验本身并不能确保一批产品无菌或已经灭菌，产品的无菌性主要通过对灭菌工艺或者无菌保障程序的验证来完成。

The test is applied to substances, preparations, or articles which, according to the Pharmacopeia, are required to be sterile. However, a satisfactory result only indicates that no contaminating microorganism has been found in the sample examined under the conditions of the test.

无菌检查法系用于检查药典要求无菌的药物、制剂产品和其他物品是否无菌的一种方法。若供试品符合无菌检查法的规定，仅表明供试品在该检验条件下未发现微生物污染。

PRECAUTIONS AGAINST MICROBIAL CONTAMINATION

预防微生物污染

The test for sterility is carried out under aseptic conditions. In order to achieve such conditions, the test environment has to be adapted to the way in which the sterility test is performed. The precautions taken to avoid contamination are such that they do not affect any microorganisms that are to be revealed in the test. The working conditions in which the tests are performed are monitored regularly by appropriate sampling of the working area and by carrying out appropriate controls.

无菌检查应在无菌条件下进行，为了达到该条件，检测环境应符合无菌检查的规定。防止污染的措施不得影响供试品中微生物的检出。检测环境应定期抽样监测并进行适当的控制。

CULTURE MEDIA AND INCUBATION TEMPERATURES

培养基和培养温度

Media for the test may be prepared as described below or equivalent commercial media may be used provided that they comply with the requirements of the Growth Promotion Test of Aerobes, Anaerobes, and Fungi.

无菌检查需制备下表所述培养基，或者是能够符合需气菌、厌氧菌、真菌促生长

试验要求的同等的商用培养基。

The following culture media have been found to be suitable for the test for sterility. Fluid Thioglycollate Medium is primarily intended for the culture of anaerobic bacteria. However, it will also detect aerobic bacteria. Soybean–Casein Digest Medium is suitable for the culture of both

fungi and aerobic bacteria.

以下培养基已经被证实适合用于无菌检查，硫乙醇酸盐流体培养基主要用于厌氧菌的培养，但其也可用于需气菌培养。大豆-酪胨培养基适合于培养真菌和需气菌。

灭菌后pH：7.1±0.2。

Mix the L-cystine, agar, sodium chloride, dextrose, yeast extract, and pancreatic digest of casein with the purified water, and heat until solution is effected. Dissolve the sodium thioglycollate or thioglycolic acid in the solution and, if necessary, add 1 N sodium hydroxide so that, after sterilization, the solution will have a pH of 7.1 ± 0.2. If filtration is necessary, heat the solution again without boiling, and filter while hot through moistened filter paper. Add the resazurin sodium solution, mix, and place the medium in suitable vessels that provide a ratio of surface to depth of medium such that not more than the upper half of the medium has undergone a color change indicative of oxygen uptake at the end of the incubation period. Sterilize using a validated process.

If the medium is stored, store at a temperature between 2 and 25 in a sterile, airtight container. If more than the upper one-third of the medium has acquired a pink color, the medium may be restored once by heating the containers in a water-bath or in free-flowing steam until the pink color disappears and by cooling quickly, taking care to prevent the introduction of nonsterile air into the container. Do not use the medium for a longer storage period than has been validated.

将L-胱氨酸、氯化钠、葡萄糖、酵母提取物、酪蛋白胰酶消化物与纯化水混合，并加热至溶解。将硫乙醇酸钠或硫乙醇酸溶解于该溶液，如果需要可加入1mol/L 氢氧化钠溶液，以便在灭菌后该溶液呈pH值7.1±0.2。如需要过滤，再次加热该溶液但不得煮沸，并趁热以湿润滤纸将该溶液过滤。加入刃天青钠溶液，混匀，并将该培养基置于适当容器中，该容器应为培养基提供特定的面积/深度比，以使在培养期结束后能明确显示氧气摄入的变色部分不超过培养基的上半部分。使用经过验证的工艺进行灭菌。如果需要贮存该培养基，应将其置于无菌、气密容器中，在2～25℃贮藏。如果超过三分之一的培养基已经呈粉红色，可以用以下方法恢复该培养基功能，但每批培养基仅能恢复一次：在水浴锅中或者自由流动蒸汽中加热该容器，直至粉色消失，并迅速放凉，须小心防止非无菌空气进入到容器中。灭菌后培养基存放时间超过验证期限时，不得使用。

Fluid Thioglycollate Medium is to be incubated at 30°–35° . For products containing a mercurial preservative that cannot be tested by the membrane filtration method, Fluid Thioglycollate Medium incubated at 20°–25° may be used instead of Soybean–Casein Digest Medium provided that it has been validated as described in Growth Promotion Test of Aerobes, Anaerobes, and Fungi. Where prescribed or justified and authorized, the following alternative thioglycollate medium might be used. Prepare a mixture having the same composition as that of the Fluid Thioglycollate Medium, but omitting the agar and the resazurin sodium solution. Sterilize as directed above. The pH after sterilization is 7.1 ± 0.2. Heat in a water bath prior to use and incubate at 30 –35 under anaerobic conditions.

硫乙醇酸盐流体培养基应在30～35℃条件下进行培养。含有汞制剂防腐剂的产品不能使用膜过滤方法检测。经需气菌、厌氧菌、真菌促生长试验验证，在20～25℃培养时，大豆酪蛋白消化培养基可以替代硫乙醇酸盐流体培养基。经合理授权，配制的与硫乙醇酸盐流体培养基成分相同，但省略了琼脂和刃天青溶液的培养基，可以替代硫乙醇酸盐流体培养基使用。按上述方法灭菌，灭菌后pH值为pH：7.1±0.2。使用前用水浴加热，置于30～35℃厌氧条件下培养。

pH after sterilization: 7.3±0.2.

Dissolve the solids in the Purified Water, heating slightly to effect a solution. Cool the solution to room temperature, and adjust the pH with 1 N sodium hydroxide so that, after sterilization, it will have a pH of 7.3 ±0.2. Filter, if necessary to clarify, dispense into suitable containers, and sterilize using a validated procedure. Store at a temperature between 2° and 25°in a sterile well-closed container, unless it is intended for immediate use. Do not use the medium for a longer storage period than has been validated.

将固体物质置纯化水中，轻微加热使其溶解。溶液放凉至室温，并用1mol/L氢氧化钠溶液调整pH值，使其灭菌后pH值为7.1±0.2。如需要使之澄清，则过滤，分装入适合的容器，并用经过验证的程序灭菌。如果不立刻使用，则保存在2～25℃无菌且密闭良好的容器中。灭菌后培养基存放时间超过验证期限时，不得使用。

Soybean–Casein Digest Medium is to be incubated at 22.5° ± 2.5°.

大豆-酪胨消化物培养基在22.5℃±2.5℃条件下培养。

◆Media for Penicillins or Cephalosporins

用于青霉素和头孢菌素的培养基

Where sterility test media are to be used in the Direct Inoculation of the Culture Medium method under Test for Sterility of the Product to be Examined, modify the preparation of Fluid Pancreatic Digest of Casein 17.0 g Papaic Digest of Soybean Meal 3.0 g Sodium Chloride 5.0 g Dibasic Potassium Phosphate 2.5 g Dextrose Monohydrate/Anhydrous 2.5/2.3 g Purified Water 1000 mL Thioglycollate Medium and the Soybean–Casein Digest Medium as follows. To the containers of each medium, transfer aseptically a quantity of -lactamase sufficient to inactivate the amount of antibiotic in the specimen under test. Determine the quantity of -lactamase required to inactivate the antibiotic by using a -lactamase preparation that has been assayed previously for its penicillin- or cephalosporin-inactivating power. [NOTE—Supplemented -lactamase media can also be used in the membrane filtration test. ]

当无菌检查培养基用于供试产品无菌检查项下的直接接种法检验时，按如下内容变更硫乙醇酸盐流体培养基和大豆-酪胨培养基的制备方法。向每一种培养基的容器中，以无菌操作转移足够量能灭活供试样品中抗生素的β-内酰胺酶（之前已经对β-内酰胺酶制品灭活青霉素或头孢菌素的能力进行过测定，据此来确定灭活抗生素所需的β-内酰胺酶量）。（注：添加β-内酰胺酶的培养基也可以用于膜过滤试验）。

Alternatively (in an area completely separate from that used for sterility testing),

confirm that an appropriate amount of -lactamase is incorporated into the medium, following either method under Method Suitability Test, using less than 100 colony-forming units (cfu) of Staphylococcus aureus (see Table 1) as the challenge. Typical microbial growth of the inoculated culture must be observed as a confirmation that the -lactamase concentration is appropriate.◆

或者（在与无菌试验完全隔离的区域），按照验证试验项下的任意一种方法，使用少于100cfu的金黄色葡萄球菌（表1）作为验证菌，来确认该培养基中含有的β-内酰胺酶量是否适宜。必须观测到接种后培养物中出现典型的微生物生长，才能确认β-内酰胺酶量是适宜的。

生物是普通拟杆菌。

The media used comply with the following tests, carried out before, or in parallel, with the test on the product to be examined.

所使用的培养基须符合下列试验，这些试验应在供试产品检验前完成或者同时进行。

Sterility

培养基无菌性

Incubate portions of the media for 14 days. No growth of microorganisms occurs.

取部分检验用培养基，连续培养14天，应不得出现微生物生长。

Growth Promotion Test of Aerobes, Anaerobes, and Fungi

需气菌、厌氧菌和真菌的促生长试验

Test each lot of ready-prepared medium and each batch of medium prepared either from dehydrated medium or from ingredients. Suitable strains of microorganisms are indicated in Table 1.

每一批配制好的培养基（采用脱水培养基或按配方制备的培养基）均需进行检查，使用的微生物菌株见表1。

Inoculate portions of Fluid Thioglycollate Medium with a small number (not more than 100 cfu) of the following microorganisms, using a separate portion of medium for each of the following species of microorganism: Clostridium sporogenes, Pseudomonas aeruginosa, and Staphylococcus aureus. Inoculate portions of alternative thioglycollate medium with a small number (not more than 100 cfu) of Clostridium sporogenes. Inoculate portions of Soybean–Casein Digest Medium with a small number (not more than 100 cfu) of the following microorganisms, using a separate portion of medium for each of the following species of microorganism: Aspergillus brasiliensis, Bacillus subtilis, and Candida albicans. Incubate for not more than 3 days in the case of bacteria and not more than 5 days in the case of fungi. Seed lot culture maintenance techniques (seed-lot systems) are used so that the viable microorganisms used for inoculation are not more than five passages removed from the original master seed-lot.

去取部分硫乙醇酸盐流体培养基，接种少量（不超过100cfu）下列微生物，每一种微生物均分别接种于单独培养基中：生孢梭菌、铜绿假单胞菌、金黄色葡萄球菌（在替代硫乙醇酸盐的流体培养基中接种少量（不超过100cfu）生孢梭菌）。在大豆-酪胨消化物培养基中接种少量（不超过100cfu）下列微生物，每一种微生物均接种于单独的培养基：黑曲霉、枯草芽孢杆菌、白色念珠菌。细菌培养时间不超过3天，真菌培养时间不超过5天。采用适宜的菌种保藏技术，以确保用于接种的微生物的传代次数不超过5代。

The media are suitable if a clearly visible growth of the microorganisms occurs.如果可见清晰的微生物生长，则该培养基是符合要求的。

DILUTING AND RINSING FLUIDS FOR MEMBRANE FILTRATION

用于膜过滤的稀释剂和冲洗液

Fluid A

稀释剂A

PREPARATION

制备

Dissolve 1 g of peptic digest of animal tissue in water to make 1 L, filter or centrifuge to clarify, if necessary, and adjust to a pH of 7.1 ± 0.2. Dispense into containers, and sterilize using a validated process.

将1g动物组织胃蛋白酶消化物溶于1L水中，如果需要则通过过滤或离心使其澄清，再调节pH值至7.1±0.2。分装入容器中，并用经过验证的工艺灭菌。

PREPARATION FOR PENICILLINS OR CEPHALOSPORINS

用于青霉素或头孢菌素的稀释剂制备

Aseptically add to the above Preparation, if necessary, a quantity of sterile -lactamase sufficient to inactivate any residual antibiotic activity on the membranes after the solution of the test specimen has been filtered (see Media for Penicillins or Cephalosporins).

在供试样品溶液已经过滤之后，如果需要，向上述制备的稀释剂中，以无菌操作加入数量足够灭活滤膜上残余抗生素的β-内酰胺酶（见用于青霉素或头孢菌素的培养基）。

Fluid D

稀释剂D

To each L of Fluid A add 1 mL of polysorbate 80, adjust to a pH of 7.1 ± 0.2, dispense into containers, and sterilize using a validated process. Use this fluid for articles containing lecithin or oil, or for devices labeled as ―sterilepathway.‖向每升稀释剂A中，加入1mL聚山梨酯80，调节pH值至7.1±0.2，分装入容器中，并使用经过验证的工艺灭菌。此液体用于含有卵磷脂或油脂的样品，或用于标为“无菌通道”的器械。

Fluid K

稀释剂K

Dissolve 5.0 g of peptic digest of animal tissue, 3.0 g of beef extract, and 10.0 g of polysorbate 80 in water to make 1 L. Adjust the pH to obtain, after sterilization, a pH of 6.9 ± 0.2. Dispense into containers, and sterilize using a validated process.

将5.0g动物组织胃蛋白酶消化物、3.0牛肉提取物、10.0g聚山梨酯80溶解于1L水中。调节pH值，使其灭菌后pH值为6.9±0.2。分装入容器中，并使用经过验证的工艺灭菌。

METHOD SUITABILITY TEST

方法适用性试验

Carry out a test as described below under Test for Sterility of the Product to be Examined using exactly the same methods, except for the following

modifications.

严格按照供试产品无菌检查项下的方法进行无菌检查。当使用到以下方法并需要进行方法调整时，需重新进行方法适用性实验。

Membrane Filtration

膜过滤法

After transferring the content of the container or containers to be tested to the membrane, add an inoculum of a small number of viable microorganisms (not more than 100 cfu) to the final portion of sterile diluent used to rinse the filter. 在将一个或多个供试容器中的内容物转移到滤膜之后，在最后一次的冲洗液中加入少量（不超过100cfu）测试菌株。

Direct Inoculation

直接接种法

After transferring the contents of the container or containers to be tested (for catgut and other surgical sutures for veterinary use: strands) to the culture medium, add an inoculum of a small number of viable microorganisms (not more than 100 cfu) to the medium.

在将一个或多个供试容器（对于兽医用的肠线和其他外科缝合用线：若干股线）中的内容物转移至培养基之后，将少量试验菌（不超过100cfu）加入培养基中。In both cases use the same microorganisms as those described above under Growth Promotion Test of Aerobes, Anaerobes, and Fungi. Perform a growth promotion test as a positive control. Incubate all the containers containing medium for not more than 5 days.

以上两种情况，均使用上述需气菌、厌氧菌、真菌促生长试验项下规定的菌株。同时设置促生长试验作为阳性对照。微生物在相应培养基中的培养时间不超过5天。

If clearly visible growth of microorganisms is obtained after the incubation, visually comparable to that in the control vessel without product, either the product possesses no antimicrobial activity under the conditions of the test or such activity has been satisfactorily eliminated. The test for sterility may then be carried out without further modification.

若培养后可见清晰的微生物生长，外观与未接种样品的对照菌生长类似，则认为该产品在此试验条件下没有抑菌作用，或者认为可能存在的抑菌作用已经被较完全的消除。此时可以认为该方法无需进一步的变更，无菌试验依法检查即可。

If clearly visible growth is not obtained in the presence of the product to be tested, visually comparable to that in the control vessels without product, the product possesses antimicrobial activity that has not been satisfactorily eliminated under the conditions of the test. Modify the conditions in order to eliminate the antimicrobial activity, and repeat the Method Suitability Test.

与未接种样品的对照相比，若在供试品存在条件下不能观察到肉眼可见的混浊，则认为该试验条件下不能消除供试品的抑菌作用。需要对实验条件进行调整以消除抗菌活性，并重新进行方法适用性试验。

This method suitability is performed (a) when the test for sterility has to be

carried out on a new product; and (b) whenever there is a change in the experimental conditions of the test. The method suitability may be performed simultaneously with the Test for Sterility of the Product to be Examined.

当一个新产品进行无菌试验时和无论何时无菌试验的试验条件发生改变时，则需进行此验证试验。该验证可以与供试产品无菌检查同时进行。

TEST FOR STERILITY OF THE PRODUCT TO BE EXAMINED

供试产品无菌检查

Number of Articles to Be Tested

供试品数量

Unless otherwise specified elsewhere in this chapter or in the individual monograph, test the number of articles specified in Table 3. If the contents of each article are of sufficient quantity (see Table 2), they may be divided so that equal appropriate portions are added to each of the specified media. [NOTE—Perform sterility testing employing two or more of the specified media. ] If each article does not contain sufficient quantities for each medium, use twice the number of articles indicated in Table 3.

除非在本章节的其他部分或在具体的各论中另有规定，供试物品的数量遵照表3中的规定。如果每个被检验物品的内容物有足够数量（见表2），可以将其分成若干等份，将适当的等份加入到每个指定的培养基。（注：使用两个或更多指定培养基，来进行无菌试验。）如果每个被检验物品的内容物规格不能满足单个培养基的接种量要求时，检验量扩大为表3所规定的检验数量的2倍。

②如果一个容器的内容物足够接种2个培养基，则此表格给出的容器数量为用于全部2个培养基的数量。

The test may be carried out using the technique of Membrane Filtration or by Direct Inoculation of the Culture Medium with the product to be examined. Appropriate negative controls are included. The technique of membrane filtration is used whenever the nature of the product permits; that is, for filterable aqueous preparations, for alcoholic or oily preparations, and for preparations miscible with, or soluble in, aqueous or oily solvents, provided these solvents do not have an antimicrobial effect in the conditions of the test.无菌检验可以使用膜过滤法或培养基直接接种法进行，应设置适宜的阴性对照。但只要该产品的性质许可，就应使用膜过滤法，也就是说，对水溶性制剂、酒精或油性制剂、易混合或溶解于水或油性溶剂的制剂，即使没有抑菌作用，也均应尽量采用薄膜过滤法进行无菌检查。

Membrane Filtration

膜过滤法

Use membrane filters having a nominal pore size not greater than 0.45 μm, in which the effectiveness to retain microorganisms has been established. Cellulose nitrate filters, for example, are used for aqueous, oily, and weakly alcoholic solutions; and cellulose acetate filters, for example, are used for strongly alcoholic solutions. Specially adapted filters may be needed for certain products (e.g., for antibiotics).

使用标称孔径不大于0.45μm的膜过滤器，此孔径已知能够有效截留微生物。例如，硝酸纤维素过滤器可用于水、油、烯醇溶液；而醋酸纤维素可用于浓醇溶液。特定产品（例如抗生素）可能需要特别改造过的特殊过滤器。

The technique described below assumes that membranes about 50 mm in diameter will be used. If filters of a different diameter are used, the volumes of the dilutions and the washings should be adjusted accordingly. The filtration apparatus and membrane are sterilized by appropriate means. The apparatus is designed so that the solution to be examined can be introduced and filtered under aseptic conditions: it permits the aseptic removal of the membrane for transfer to the medium, or it is suitable for carrying out the incubation after adding the medium to the apparatus itself.

以下涉及的方法中所使用的均为直径约50mm的滤膜。如果使用不同直径的过滤器，稀释液和冲洗液的体积应当做相应调节。过滤器和滤膜都应该经过灭菌处理。过滤器应该满足在无菌条件下可以灌注并过滤供试溶液的要求：使得在无菌状态下摘掉滤膜，并将其转移至培养基中成为可能；或者应满足将培养基灌注至该设备中，进行培养的要求。

AQUEOUS SOLUTIONS

水溶性溶液

If appropriate, transfer a small quantity of a suitable, sterile diluent such as Fluid A (see Diluting and Rinsing Fluids for Membrane Filtration) onto the membrane in the apparatus and filter. The diluent may contain suitable neutralizing substances and/or appropriate inactivating substances, for example, in the case of antibiotics.

应当尽量减少转移至过滤器滤膜上的无菌稀释剂，例如稀释剂A（见用于膜过滤的稀释剂和冲洗液）。稀释剂中可能会含有一定量的中和物质或灭活物质，例如对抗生素的检测。

Transfer the contents of the container or containers to be tested to the membrane or membranes, if necessary, after diluting to the volume used in the Method Suitability Test with the chosen sterile diluent, but using not less than the quantities of the product to be examined prescribed in Tables 2 and 3. Filter immediately. If the product has antimicrobial properties, wash the membrane not less than three times by filtering through it each time the volume of the chosen sterile diluent used in the Method Suitability Test. Do not exceed a washing cycle of five times 100 mL per filter, even if during method suitability it has been demonstrated that such a cycle does not fully eliminate the antimicrobial activity. Transfer the whole membrane to the culture medium or cut it aseptically into two equal parts, and transfer one half to each of two suitable media. Use the same volume of each medium as in the Method Suitability Test. Alternatively, transfer the medium onto the membrane in the apparatus. Incubate the media for not less than 14 days.

将一个或多个供试容器的内容物转移到滤膜，并立即滤过。如需要可先用选定的无菌稀释剂稀释至方法适用性试验中所用的体积，但须使用不少于表2和表3中规定的供试品数量。如果该产品具有抗菌活性，滤膜至少冲洗3次，每次冲洗量均按照方法适用性试验中确定的无菌稀释剂体积冲洗滤膜。冲洗过程通常不应该超过“5次，100mL/次”的冲洗循环，即便该冲洗循环不能完全消除抗菌活性。将整个滤膜转移至培养基，或以无菌操作将滤膜切至相等的2部分，并将每

一部分转移至适当的培养基中。每个培养基的体积按照方法适用性试验所确定的用量。或者，将培养基转移至带有滤膜的滤器中。培养该培养基，不少于14天。

SOLUBLE SOLIDS

可溶性固体

Use for each medium not less than the quantity prescribed in Tables 2 and 3 of the product dissolved in a suitable solvent, such as the solvent provided with the preparation, Sterile Water for Injection, sterile saline, or a suitable sterile solution such as Fluid A (Diluting and Rinsing Fluids for Membrane Filtration), and proceed with the test as described above for Aqueous Solutions using a membrane appropriate to the chosen solvent.

在每个培养基中，使用不少于表2和表3规定的产品数量溶于适当溶剂，例如稀释剂A（用于膜过滤的稀释剂和冲洗液），并按照上述关于水性溶液样品实验操作的要求，使用适合所选溶液的滤膜，进行试验。

OILS AND OILY SOLUTIONS

油和油性溶液

Use for each medium not less than the quantity of the product prescribed in Tables 2 and 3. Oils and oily solutions of sufficiently low viscosity may be filtered without dilution through a dry membrane. Viscous oils may be diluted as necessary with a suitable sterile diluent such as isopropyl myristate shown not to have antimicrobial activity in the conditions of the test. Allow the oil to penetrate the membrane by its own weight, and then filter, applying the pressure or suction gradually. Wash the membrane at least three times by filtering through it each time about 100 mL of a suitable sterile solution such as Fluid A (see Diluting and Rinsing Fluids for Membrane Filtration) containing a suitable emulsifying agent at a concentration shown to be appropriate in

the Method Suitability Test, for example polysorbate 80 at a concentration of 10 g per L (Fluid K) . Transfer the membrane or membranes to the culture medium or media, or vice versa, as described above for Aqueous Solutions, and incubate at the same temperatures and for the same times.

在每个培养基中，使用不少于表2和表3中规定的检验量。黏性较低的油和油性溶液可在不经稀释的情况下滤过干燥滤膜。黏稠油质可以用适合的无菌稀释剂进行稀释，如已证实在该试验条件下不具有抗菌活性的豆蔻酸异丙酯。油质依靠其自身的重量穿过滤膜，然后逐渐加压或抽吸过滤。使用适宜的无菌冲洗液冲洗滤膜，至少冲洗3次，约100mL/次，例如使用一定浓度乳化剂的稀释剂A（参见用于膜过滤的稀释剂和冲洗液）。通过方法适用性试验确定乳化剂的适宜浓度，例如浓度为每升10克的聚山梨酯80（稀释剂K）。将一个或多个滤膜转移到一个或多个培养基；或者也可以参照关于水性溶液的相关描述进行操作，将培养基在上述规定的温度下培养相同时间。

OINTMENTS AND CREAMS

油膏和乳膏

Use for each medium not less than the quantities of the product prescribed in

Tables 2 and 3. Ointments in a fatty base and emulsions of the water-in-oil type may be diluted to 1% in isopropyl myristate as described above, by heating, if necessary, to not more than 40°. In exceptional cases it may be necessary to heat to not more than 44°. Filter as rapidly as possible, and proceed as described above for Oils and Oily Solutions.

在每个培养基中，使用不少于表2和表3中规定的样品量。参照上文，脂肪状的油膏和油包水的乳液可用豆蔻酸异丙酯稀释至1%浓度，如需要可加热，但不超过40℃。在特殊情况下，加热温度可适当增加但不超过44℃。供试液制备好后尽快过滤，并参照油和油性溶液的要求进行试验。

PREFILLED SYRINGES

预装填的注射器

For prefilled syringes without attached sterile needles, expel the contents of each syringe into one or two separate membrane filter funnels or into separate pooling vessels prior to transfer. If a separate sterile needle is attached, directly expel the syringe contents as indicated above, and proceed as directed for Aqueous Solutions. Test the sterility of the needle, using Direct Inoculation under Method Suitability Test.

对于没有附无菌针头的预装填注射器，在转移之前，将每个注射器的内容物排出一个或两个单独的膜过滤器漏斗，或独立并联的容器。如果附有单独的灭菌针头，直接按照上面的规定将注射器内容物直接排出，并按照关于水性溶液的规定进行试验。参照方法适用性试验项下的直接接种法，检查针头的无菌情况。

SOLIDS FOR INJECTION OTHER THAN ANTIBIOTICS

非抗生素注射用固体

Constitute the test articles as directed on the label, and proceed as directed for Aqueous Solutions or Oils and Oily Solutions, whichever applies. [NOTE—If necessary, excess diluent can be added to aid in the constitution and filtration of the constituted test article. ]

按照其标签上的规定配制供试物品，并按照适用的关于水性溶液或油和油性溶液的规定继续进行。（注：如需要，可以加入额外的稀释剂以帮助对已配制的供试物品进行再配制和过滤）。

ANTIBIOTIC SOLIDS FOR INJECTION

注射用抗生素固体

Pharmacy Bulk Packages, <5 g—From each of 20 containers, aseptically transfer about 300 mg of solids, into a sterile 500-mL conical flask, dissolve in about 200 mL of Fluid A (see Diluting and Rinsing Fluids for Membrane Filtration), and mix; or constitute, as directed in the labeling, each of 20 containers and transfer a quantity of liquid or suspension, equivalent to about 300 mg of solids, into a sterile 500-mL conical flask, dissolve in about 200 mL of Fluid A, and mix. Proceed as directed for Aqueous Solutions or Oils and Oily Solutions, whichever applies.

原料包装，＜5g 取20个容器，无菌操作从每个容器中取约300mg固体至500mL

无菌锥形瓶，使用约200mL稀释剂A溶解并混匀（参见用于膜过滤的稀释剂和冲洗液），或取20个容器，每个容器均按照标签上的规定配制，并从每个容器中将相当于300mg固体的液体转移至500mL无菌锥形瓶，使用约200mL稀释剂A溶解并混匀。按照适合的关于水性溶液或油和油性溶液的规定，继续操作。Pharmacy Bulk Packages, ≥5 g—From each of 6 containers, aseptically transfer about 1 g of solids into a sterile 500-mL conical flask, dissolve in about 200 mL of Fluid A, and mix; or constitute, as directed in the labeling, each of 6 containers and transfer a quantity of liquid, equivalent to about 1 g of solids, into a sterile 500-mL conical flask, dissolve in about 200 mL of Fluid A, and mix. Proceed as directed for Aqueous Solutions.

原料包装，≥5g 取6个容器，无菌操作从每个容器中取约1g固体转移至500mL 无菌锥形瓶，使用约200mL稀释剂A溶解并混匀；或取6个容器，每个均按照标签的规定配制，将相当于1g固体的液体转移至500mL无菌锥形瓶，使用约200mL 稀释剂A溶解并混匀。按照水性溶液的规定，继续操作。

ANTIBIOTIC SOLIDS, BULKS, AND BLENDS

抗生素固体、原料及预混物

Aseptically remove a sufficient quantity of solids from the appropriate amount of containers (see Table 2), mix to obtain a composite, equivalent to about 6 g of solids, and transfer to a sterile 500-mL conical flask. Dissolve in about 200 mL of Fluid A, and mix. Proceed as directed for Aqueous Solutions.

无菌操作从适当数量的容器中（见表2）分别取出足够数量的固体，混匀得6g 固体的混合物，至500mL无菌锥形瓶。使用约200mL稀释剂A溶解并混匀。按照水性溶液的规定，继续操作。

STERILE AEROSOL PRODUCTS

无菌气（喷）雾剂产品

For fluid products in pressurized aerosol form, freeze the containers in an alcohol-dry ice mixture at least at –20 for about 1 hour. If feasible, allow the propellant to escape before aseptically opening the container, and transfer the contents to a sterile pooling vessel. Add 100 mL of Fluid D to the pooling vessel, and mix gently. Proceed as directed for Aqueous Solutions or Oils and Oily Solutions, whichever applies.

对于以加压气（喷）雾剂形式存在的液体产品，在大约－20℃的“酒精-干冰”混合物中冷冻约1小时。若可行，在以无菌操作打开容器之前去除抛射剂，并将内容物转移至一个无菌的容器。向容器中加入100mL稀释剂D，并轻轻混匀。并根据情况，按照水性溶液或油和油性溶液的规定继续操作。

DEVICES WITH PATHWAYS LABELED STERILE

标示为无菌的具有管道的医疗器械

Aseptically pass not less than 10 pathway volumes of Fluid D through each device tested. Collect the fluids in an appropriate sterile vessel, and proceed as directed for Aqueous Solutions or Oils and Oily Solutions, whichever applies.

以无菌操作使用稀释剂D冲洗每个器械样本，并且冲洗量不少于器械管道体积的

10倍。在适当的无菌容器中收集冲洗液，并根据情况，按照水性溶液或油和油性溶液的规定继续操作。

In the case of sterile, empty syringes, draw sterile diluent into the barrel through the sterile needle, if attached, or through a sterile needle attached for the purpose of the test, and express the contents into a sterile pooling vessel. Proceed as directed above.

对于无菌的空注射器，如果附有无菌针头，可直接将无菌稀释剂吸取至注射器，或者使用一个专为此试验准备的无菌针头吸取稀释剂，并将内容物压出至合并容器中。按照上述内容继续进行。

Direct Inoculation of the Culture Medium

培养基的直接接种法

Transfer the quantity of the preparation to be examined prescribed in Tables 2 and 3 directly into the culture medium so that the volume of the product is not more than 10% of the volume of the medium, unless otherwise prescribed.

按照表2和表3规定的供试品取样量，将该供试品直接转移到培养基中，除另有规定外，接种量不得超过培养基体积的10%。

If the product to be examined has antimicrobial activity, carry out the test after neutralizing this with a suitable neutralizing substance or by dilution in a sufficient quantity of culture medium. When it is necessary to use a large volume of the product, it may be preferable to use a concentrated culture medium prepared in such a way that it takes into account the subsequent dilution. Where appropriate, the concentrated medium may be added directly to the product in its container.

如果供试品具有抗菌活性，使用适宜的中和剂或采用培养基稀释法消除其抗菌活性后，再进行该试验。当供试品体积较大时，最后配制使用浓缩培养基，以便后续的稀释需要。若条件适宜，浓缩培养基可以直接加入到装有产品的容器中。

OILY LIQUIDS

油性液体

Use media to which have been added a suitable emulsifying agent at a concentration shown to be appropriate in the Method Suitability Test, for example polysorbate 80 at a concentration of 10 g per L.

使用已经加入了适当乳化剂的培养基，乳化剂浓度需经方法适用性试验证明适用于该试验，例如浓度为每升10g的聚山梨酯80。

OINTMENTS AND CREAMS

油膏和乳膏

Prepare by diluting to about 1 in 10 by emulsifying with the chosen emulsifying agent in a suitable sterile diluent such as Fluid A (see Diluting and Rinsing Fluids for Membrane Filtration). Transfer the diluted product to a medium not containing an emulsifying agent.

按照1:10比例将选定的乳化剂混匀在适当的无菌稀释液中，例如稀释剂A（见用于膜过滤的稀释剂和冲洗液），以含有乳化剂的稀释液来制备供试液。将供试

液转移至不含乳化剂的培养基中。

Incubate the inoculated media for not less than 14 days. Observe the cultures several times during the incubation period. Shake cultures containing oily products gently each day. However, when Fluid Thioglycollate Medium is used for the detection of anaerobic microorganisms, keep shaking or mixing to a minimum in order to maintain anaerobic conditions.

接种后的培养基培养不少于14天。在培养中隔日观察。每天轻轻摇动含有油性产品的培养基。但是，当使用硫乙醇酸盐流体培养基用于检测厌氧微生物培养基时，尽量减少摇动，以维持厌氧条件。

CATGUT AND OTHER SURGICAL SUTURES FOR VETERINARIAN USE

兽医用肠线和其他外科缝合线

Use for each medium not less than the quantities of the product prescribed in Tables 2 and 3. Open the sealed package using aseptic precautions, and remove three sections of the strand for each culture medium. Carry out the test on three sections, each 30-cm long, which have been cut off from the beginning, the center, and the end of the strand. Use whole strands from freshly opened cassette packs. Transfer each section of the strand to the selected medium. Use sufficient medium to cover adequately the material to be tested (20 mL to 150 mL).

在每个培养基中，使用不少于表2和表3中所规定的检验品。无菌操作打开封闭的包装，使用从刚刚打开的包装盒取出整股线，从该线的前端、中间、末端分别截取每段30cm的产品进行试验，每种培养基均接种由该股线截取的3部分供试物料。使用充足的培养基，以充分覆盖供试物料（20～150mL）。

SOLIDS

固体

Transfer a quantity of the product in the form of a dry solid (or prepare a suspension of the product by adding sterile diluent to the immediate container), corresponding to not less than the quantity indicated in Tables 2 and 3. Transfer the material so obtained to 200 mL of Fluid Thioglycollate Medium, and mix. Similarly, transfer the same quantity to 200 mL of Soybean–Casein Digest Medium, and mix. Proceed as directed above.

转移干燥固体产品（或通过加入无菌稀释剂在中间容器中配制的该产品的悬浮液），检验量不少于表2和表3中的规定。将供试品接种至200mL硫乙醇酸盐流体培养基中，混匀。同法接种至200mL大豆-酪蛋白消化物培养基中，混匀。按照上述规定继续操作。

PURIFIED COTTON, GAUZE, SURGICAL DRESSINGS, AND RELATED ARTICLES

脱脂棉花、纱布、外科敷料及相关物品

From each package of cotton, rolled gauze bandage, or large surgical dressings being tested, aseptically remove two or more portions of 100- to 500-mg each from the innermost part of the sample. From individually packaged, single-use materials, aseptically remove the entire article. Immerse the portions or article in each medium, and proceed as directed above.

从待检的每个包装中的棉花、卷状纱布绷带、大块外科敷料中，以无菌操作从中心部位取出2个或更多份样品，每份样品100～500mg。对独立包装的一次性使用样品，以无菌操作取出整个样品。将这些取样样品或整个样品浸没在各个培养基中，继续按照上述内容操作。

STERILE DEVICES

无菌器械

Articles can be immersed intact or disassembled. To ensure that device pathways are also in contact with the media, immerse the appropriate number of units per medium in a volume of medium sufficient to immerse the device completely, and proceed as directed above. For extremely large devices, immerse those portions of the device that are to come into contact with the patient in a volume of medium sufficient to achieve complete immersion of those portions.

样品可以完整地拆开后浸没在培养中。为确保器械管道与培养基接触，使用足够体积的培养基浸没整个器械零部件，并按照上述内容继续操作。对于极大的器械，将该器械中将要与患者接触的部分，浸没到足够体积的培养基中。

For catheters where the inside lumen and outside are required to be sterile, either cut them into pieces such that the medium is in contact with the entire lumen or fill the lumen with medium, and then immerse the intact unit.

对于管道内部的腔体和外部均要求无菌的导尿管，将它们切片，这样培养基就可以接触整个腔体，或者用培养基填充腔体，然后浸没整个器械。

OBSERVATION AND INTERPRETATION OF RESULTS

结果的观察和判断

At intervals during the incubation period and at its conclusion, examine the media for macroscopic evidence of microbial growth. If the material being tested renders the medium turbid so that the presence or absence of microbial growth cannot be readily determined by visual examination, 14 days after the beginning of incubation transfer portions (each not less than 1 mL) of the medium to fresh vessels of the same medium, and then incubate the original and transfer vessels for not less than 4 days.

无菌检查在培养观察期间和下结论时，都要检查该培养基是否有肉眼可见的微生物生长。如果供试物料导致培养基混浊，但无法通过肉眼观察来确定是否存在微生物生长时，培养14天之后，将该培养基的若干部分（每个部分不少于1mL）转接种至含相同新鲜培养基的容器中，培养不少于4天。

If no evidence of microbial growth is found, the product to be examined complies with the test for sterility. If evidence of microbial growth is found, the product to be examined does not comply with the test for sterility, unless it can be clearly demonstrated that the test was invalid for causes unrelated to the product to be examined. The test may be considered invalid only if one or more of the following conditions are fulfilled:

如果未发现微生物生长，则认为该供试产品无菌检查结果合格。若发现微生物生长，除非有充分证据证实此次试验无效或微生物污染与该供试产品无关，否则认

为该供试产品无菌检查不合格。当满足下一个或多个条件时，认为检验结果阳性无效。

a. The data of the microbiological monitoring of the sterility testing facility show

a fault.

b. A review of the testing procedure used during the test in question reveals a fault.

c. Microbial growth is found in the negative controls.

d. After determination of the identity of the microorganisms isolated from the test, the growth of this species (or these species) may be ascribed unequivocally to faults with respect to the material and or the technique used in conducting the sterility test procedur

e.

①该无菌试验设施的微生物监控数据显示实验保障有缺陷。

②通过实验回顾发现实验过程中有疑问提示有实验过程缺陷。

③阳性对照中发现微生物生长。

④阳性结果中的微生物经过菌种鉴别后，确定该（或这些）微生物的生长与实验材料无关和/或与无菌检查方法有关。

If the test is declared to be invalid, it is repeated with the same number of units as in the original test. If no evidence of microbial growth is found in the repeat test, the product examined complies with the test for sterility. If microbial growth is found in the repeat test, the product examined does not comply with the test for sterility.

如果该试验证明无效，应取用与原试验相同量的供试品进行重验。若在重新试验中未发现微生物生长，则认为该产品符合无菌要求。若在重验中发现微生物生长，则该产品不合格。

APPLICATION OF THE TEST TO PARENTERAL PREPARATIONS,

OPHTHALMIC, AND OTHER NONINJECTABLE PREPARATIONS REQUIRED TO COMPLY WITH THE TEST FOR STERILITY 无菌检查在注射药品、眼科和其他必须符合无菌要求的非注射药品中的应用When using the technique of membrane filtration, use, whenever possible, the whole contents of the container, but not less than the quantities indicated in Table 2, diluting where necessary to about 100 mL with a suitable sterile solution, such as Fluid A (see Diluting and Rinsing Fluids for Membrane Filtration).

当使用膜过滤法时，只要可能，就要使用包装容器的全部内容物，但不少于表2中规定的检验量，必要时以适当的无菌溶液将供试品稀释至约100mL，例如液体A（参见用于膜过滤的稀释剂和冲洗液）。

When using the technique of direct inoculation of media, use the quantities shown in Table 2, unless otherwise justified and authorized. The tests for bacterial and fungal sterility are carried out on the same sample of the product to be examined. When the volume or the quantity in a single container is insufficient to carry out the tests, the contents of two or more containers are used to inoculate the different media.

当使用直接接种法时，除非另有证据或授权，使用表2中规定的检验量。可用同