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食品中的大肠菌群和大肠杆菌的检测 991.14食品中的大肠菌群和大肠杆菌的检测再水化干膜法AOAC 991.14 再水化干膜法 再水化干膜法

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AOAC991.14食品中的大肠菌群和大肠杆菌的检测再水化干膜法（中英文版）.rar,食品中的大肠菌群和大肠杆菌的检测,991.14食品中的大肠菌群和大肠杆菌的检测再水化干膜法AOAC,991.14,再水化干膜法 ,再水化干膜法

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17.3.04 AOAC 官方方法官方方法 991.14 食品中的大肠菌群和大肠杆菌的检测食品中的大肠菌群和大肠杆菌的检测 再水化干膜法再水化干膜法 （Petrifilm 大肠菌群测试片和大肠菌群测试片和 Petrifilm 大肠杆菌测试片）大肠杆菌测试片） 首次发布首次发布 1991 最终发布最终发布 1994 方法性能方法性能 方法性能数据见表格 991.14（见下页） 。 A. 原理原理 见 989.10A（见 17.3.03） B. 设备和试剂设备和试剂 见 989.10B（b）-（f） （见 17.3.03） 大肠杆菌测试片和大肠菌群测试片相似，989.10B（b） （见 17.3.03） ，加入了 D溴-4-氯吲哚D葡萄糖苷酶，这是一种用以指示葡萄糖苷酶的试剂，这样就能在同一个测试片上同时得出大肠菌群和大肠杆菌数。Petrifilm 大肠杆菌测试片符合这些规格（生产商为 Medical Surgical Division/3M, 275-W 3M Center，St. Paul, MN 55144） 。 C. 样品制样品制备备 见 966.23B（见 17.2.01）和 986.33C（见17.3.02）. D. 分析分析 （a）大肠菌群计数将大肠杆菌测试片和大肠菌群测试片 989.10B（b） （见 17.3.03）置于平坦表面。提起上层膜，将 1mL 样品均液接种于测试片中央。小心将上层膜覆盖在接种面上。对压板（平面朝下）的中心轻轻施压，将样品分布于限定的生长区域内。将测试片静置 1 分钟，以使凝胶固化。将测试片置于 35C 的条件下培养 242小时。 将测试片水平朝上放置于培养器中。每叠不要超过 20 片。在培养结束后及时进行计数。培养结束时，测试片能够在冷冻条件下保存 7 天（-15C） ，但应避免将此作为常规操作。使用标准菌群计数器计数，也可以使用放大镜辅助计数。大肠菌群菌群含有一个或多个气泡（在一个菌落直径之内） 。 在合理计数范围(15-150 个)内计算所有的菌群数。没有气泡的红色菌群不记作大肠菌群。 （b）大肠杆菌计数使用大肠杆菌测试片，并用与（a）相同的方法处理。再多培养 242 小时（总计 484 小时 ） 。大肠杆菌为含有气泡的蓝色菌群，其它非大肠杆菌的大肠菌群为带气泡的红色菌群。 参考文献：JAOAC 74,635（1991） 修改日期：1998.3 17.3.04AOAC Official Method 991.14Coliform andEscherichia coliCounts in FoodsDry Rehydratable Film(Petrifilm?E. coli/Coliform Count Plate? andPetrifilm? Coliform Count Plate?) MethodsFirst Action 1991Final Action 1994See Table 991.14 for the results of the interlaboratory study sup-porting the acceptance of the method.A. PrincipleSee 989.10A (see 17.3.03).B. Apparatus and ReagentSee 989.10B(b)(f) (see 17.3.03).E. coli count plates.Plates are similar to coliform count plates,989.10B(b)(see17.3.03),withadditionof5-bromo-4-chloro-3-indolyl-D-glucuronide, an indicator of glucuronidase activity. This allows coliformsandE.colitobereadonthesameplate.PetrifilmE.coliCountPlates(Micro-biologyProducts,3MCenter,Bldg.275-5W-05,St.Paul,MN55144,USA)meet thesespecifications.C. Preparation of Test SuspensionSee 966.23B (see 17.2.01) and 986.33C (see 17.3.02).D. Analysis(a) Coliform count.Place dry-film E. coli count plate, B, orcoliform count plate, 989.10B(b) (see 17.3.03), on flat surface. Lifttopfilmandinoculate1mLtestsuspensionontocenteroffilmbase.Carefullyplacetopfilmdownontoinoculum.Distributetestsuspen-sionoverprescribedgrowthareawithdownwardpressureoncenterof plastic spreader device (flat side down). Leave plate undisturbed1mintopermitgeltosolidify.Incubateplates242hat351C.In incubator, place plates in horizontal position, clear side up, instacks not exceeding 20 units. Count plates promptly after incuba-tion period. After incubation is complete, plates may be storedfrozen (15C) up to 7 days. This should be avoided as routinepractice. Use standard colony counter for counting purposes. Magni-fier-illuminator may also be used to facilitate counting. Coliforms ap-pear as red colonies that have one or more gas bubbles associated(withinonecolonydiameter)withthem.Countallcoloniesincountablerange (15150 colonies). Red colonies without gas bubbles are notcounted as coliform organisms.(b) E.colicount.UseE.colicountplateandproceedasin(a). In-cubateanadditional242h(484htotal).E.colicoloniesappearasbluecoloniesassociatedwithgasbubbles;othercoliformsappearasredcolonies with gas.Reference: JAOAC 74, 635(1991).Revised: March 2002 2002 AOAC INTERNATIONAL 2002 AOAC INTERNATIONALTable 991.14Interlaboratory study results for total coliforms andE. colion dry-film count platesProductLevelMean log, cfu/gsrsRRSDr, %RSDR, %Total coliforms on coliform count platesGround turkeyLow1.8450.1930.29610.516.1Medium2.2350.3610.36116.216.2High2.7440.1050.2703.89.9Fresh mushroomsLow4.2410.2780.8086.619.0Medium4.9990.6621.14113.222.8High5.1970.1200.8722.316.8Beef with gravyLow1.0690.2430.28822.727.0Medium2.0620.2570.39112.518.9High2.7320.2330.3208.511.7CheeseLow0.7830.1720.17221.921.9Medium2.4110.0920.2063.88.5High3.4340.2120.3646.210.6Wheat flourLow2.6350.2910.87411.133.2Medium3.4600.7461.29121.637.3High4.0490.8731.55021.638.3Nut mealLow0.8960.3360.44337.549.5Medium1.3450.4860.69936.252.0High2.3700.2670.83111.335.1Total coliforms onE. colicount platesGround turkeyLow1.9340.1770.2619.213.5Medium2.2160.2850.31512.914.2High2.8350.1130.2424.08.5Fresh mushroomsLow4.5500.0790.7221.715.9Medium5.2780.6001.00111.419.0High5.2270.3460.8176.615.6Beef with gravyLow1.0160.2150.28421.227.9Medium2.0600.2560.29512.414.3High2.6790.1910.3317.112.4CheeseLow0.8130.1720.19221.123.6Medium2.5070.1090.2384.39.5High3.6200.0740.1062.12.9Wheat flourLow2.6390.2430.8839.233.4Medium3.4320.8261.31824.138.4High4.0640.9291.55722.938.8Nut mealLow0.7590.0280.2953.738.8Medium1.4030.3780.72826.951.8High2.3550.4250.91518.038.9TotalE. colionE. colicount platesGround turkeyLow1.4190.3160.31622.322.3Medium1.9230.3640.36418.918.9High2.6070.0980.2333.88.9Fresh mushroomsLow2.5000.2610.34310.413.7Medium3.0580.2960.4229.713.8High3.4360.7150.76020.822.1Beef with gravyLow0.7530.0280.1723.722.8Medium0.9910.1540.60115.660.7High1.3120.1610.82812.363.1CheeseLow0.