细菌内毒素检测中文翻译

85细菌内毒素检测PortionsofthisgeneralchapterhavebeenharmonizedwiththecorrespondingtextsoftheEuropeanPharmacopoeiaand/ortheJapanesePharmacopoeia.Thoseportionsthatarenotharmonizedaremarkedwithsymbolstospecifythisfact.本文中部分内容与欧洲药典和/或日本药典的相应的内容是一致的，那些不一致的内容则用符号（）标明TheBacterialEndotoxinsTest(BET)isatesttodetectorquantifyendotoxinsfromGram-negativebacteriausingamoebocytelysatefromthehorseshoecrab(LimuluspolyphemusorTachypleustridentatus).细菌内毒素测试(BET)是检测或定量革兰氏阴性细菌内毒素的试验，它使用从鲎（美洲鲎或东方鲎）的循环阿米巴细胞的抽提液中制备的鲎试剂。Therearethreetechniquesforthistest:thegel-clottechnique,whichisbasedongelformation;theturbidimetrictechnique,basedonthedevelopmentofturbidityaftercleavageofanendogenoussubstrate;andthechromogenictechnique,basedonthedevelopmentofcoloraftercleavageofasyntheticpeptide-chromogencomplex.Proceedbyanyofthethreetechniquesforthetest.Intheeventofdoubtordispute,thefinaldecisionismadebaseduponthegel-clot■limittest■2s(USP35)unlessotherwiseindicatedinthemonographfortheproductbeingtested.Thetestiscarriedoutinamannerthatavoidsendotoxincontamination.细菌内毒素试验有三种技术：基于凝胶形成的凝胶法；基于一内源性基质断裂后的浊度变化；和显色基质法——基于一个带有生色团的合成肽断裂后颜色的变化。除非文中另有说明，否则可以用三种技术中的任何一种进行测试。若检测结果存在争议，除非文中另有说明，以凝胶法结果为准。该试验以一种避免内毒素污染的方式进行。APPARATUS仪器设备Depyrogenateallglasswareandotherheat-stablematerialsinahotairovenusingavalidatedprocess.◆1◆Acommonlyusedminimumtimeandtemperatureis30minat250°.Ifemployingplasticapparatus,suchasmicroplatesandpipettipsforautomaticpipetters,useapparatusthatisshowntobefreeofdetectableendotoxinanddoesnotinterfereinthetest.[NOTE—Inthischapter,theterm“tube”includesanyotherreceptaclesuchasamicrotiterwell.]所有玻璃用具和耐热材料应置于热空气烤箱中有效除热原。通常使用的最少时间和最低温度为30分钟，250℃。如果使用塑料用具，如微孔板和自动加样器配套的吸头之类的，应确认其不含可测内毒素并不会干扰试验。[注：在这一章中，“管”这个词泛指其它任何容器，如微量孔板的孔]REAGENTSANDTESTSOLUTIONS试剂和测试方法AmoebocyteLysate—Alyophilizedproductobtainedfromthelysateofamoebocytes(whitebloodcells)fromthehorseshoecrab(LimuluspolyphemusorTachypleusSolutridentatus).Thisreagentrefersonlytoaproductmanufacturedinaccordancewiththeregulationsofthecompetentauthority.[NOTE—AmoebocyteLysatereactstosomeΒ-glucansinadditiontoendotoxins.AmoebocyteLysatepreparationsthatdonotreacttoglucansareavailable:theyarepreparedbyremovingtheGfactorreactingtoglucansfromAmoebocyteLysateorbyinhibitingtheGfactorreactingsystemofAmoebocyteLysateandmaybeusedforendotoxintestinginthepresenceofglucans.]鲎试剂是一种冻干的产物，它从鲎（美洲鲎或东方鲎）的阿米巴细胞（白血细胞）的溶解产物中获得。本试剂仅指根据主管当局的规定而生产的产品。[注：除了内毒素，鲎试剂也与某些β-葡聚糖反应。一些经过处理而制备的鲎试剂不会与β-葡聚糖反应，它们是通过除去阿米巴细胞产生的G因子或抑制阿米巴细胞反应系统G因子的反应机制实现的，对于含有葡聚糖的样品的检测应用此类试剂。]WaterforBacterialEndotoxinsTest(BET)—UseWaterforInjectionorwaterproducedbyotherproceduresthatshowsnoreactionwiththelysateemployed,atthedetectionlimitofthereagent.用于细菌内毒素测试的水(注):在试剂的检测范围内，使用注射用水或不含与鲎试剂反应的物质的水。LysateTS—DissolveAmoebocyteLysateinWaterforBET,orinabufferrecommendedbythelysatemanufacturer,bygentlestirring.Storethereconstitutedlysate,refrigeratedorfrozen,accordingtothespecificationsofthemanufacturer.将鲎试剂溶解于水中，或在制造商推荐的缓冲液中温和搅拌，然后根据说明书将重组的鲎试剂冷藏或冷冻。◆1Foravaliditytestoftheprocedureforinactivatingendotoxins,seeDry-HeatSterilizationunderSterilizationandSterilityAssuranceofCompendialArticles1211.UseLysateTShavingasensitivityofnotlessthan0.15EndotoxinUnitpermL.◆为使内毒素进行有效的检验，选择干热灭菌，并保证遵照1211条例的规定。使用的是一种敏感性每毫升不少于0.15个内毒素单位的鲎试剂。PREPARATIONOFSOLUTIONS准备的解决方案StandardEndotoxinStockSolution—AStandardEndotoxinStockSolutionispreparedfromaUSPEndotoxinReferenceStandardthathasbeencalibratedtothecurrentWHOInternationalStandardforEndotoxin.FollowthespecificationsinthepackageleafletandonthelabelforpreparationandstorageoftheStandardEndotoxinStockSolution.EndotoxinisexpressedinEndotoxinUnits(EU).[NOTE—OneUSPEndotoxinUnit(EU)isequaltooneInternationalUnit(IU)ofendotoxin.]标准内毒素溶液是按USP内毒素参考标准制备的，该标准已相当于目前国际内毒素的标准。按照标准内毒素溶液说明书和和储存标签的说明，内毒素的量用USP内毒素单位（USP-EU）表示。注1:一种USP内毒素单位(EU)等于一国际单位(国际单位)内毒素。[注：1USP-EU内毒素等于国际单位1IU的内毒素]StandardEndotoxinSolutions—AftermixingtheStandardEndotoxinStockSolutionvigorously,prepareappropriateserialdilutionsofStandardEndotoxinSolution,usingWaterforBET.Usedilutionsassoonaspossibletoavoidlossofactivitybyadsorption.标准内毒素溶液：在将标准内毒素溶液混合后，将标准内毒素溶液进行适当的连续稀释。尽可能使用稀释剂，以避免因吸附而失去活性。SampleSolutions—PreparetheSampleSolutionsbydissolvingordilutingdrugs■■2S(USP35)usingWaterforBET.Somesubstancesorpreparationsmaybemoreappropriatelydissolved,■ordiluted■2s(usp35)inotheraqueoussolutions.Ifnecessary,adjustthepHofthesolutiontobeexamined(ordilutionthereof)sothatthepHofthemixtureofthelysateandSampleSolutionfallswithinthepHrangespecifiedbythelysatemanufacturer,usually6.0–8.0.ThepHmaybeadjustedbyuseofanacid,base,orsuitablebufferasrecommendedbythelysatemanufacturer.AcidsandbasesmaybepreparedfromconcentratesorsolidswithWaterforBETincontainersfreeofdetectableendotoxin.Buffersmustbevalidatedtobefreeofdetectableendotoxinandinterferingfactors.供试品溶液的制备——用细菌内毒素检查用水溶解或稀释药品或抽提医疗器械来制备供试品溶液。某些物质或产品用其它水性溶液溶解、稀释或抽提可能更合适。如果有必要，调节待检溶液（或稀释液）的pH值以使鲎试剂和供试品的混合物的pH值在鲎试剂生产商指定的pH值范围内。这通常适用于pH值为6.0-8.0之间的产品。可用酸、碱或鲎试剂生产商推荐的合适的缓冲液调节pH值，酸和碱可用原液或固体物加水于无内毒素的容器中进行制备用于细菌内毒素检查。缓冲液必须经过验证确认无可测的内毒素和干扰因子。DETERMINATIONOF