Validation of the Delvotest SP NT DA. Performance Tested Method 011101

Delvotest SP NT DA is designed to test milk for the presence of antibacterial substances, such as antibiotics. The test is made of an agar gel containing bacterial spores and a color indicator. The milk sample is added onto the agar gel, and the test is incubated at 64 degrees C. The principle of the test is based on the diffusion of possible inhibitory substances that may be present in the milk sample into agar. This reduces growth and acid production by the test organism, and delays or prevents the agar from changing color from purple to yellow. The Delvotest Accelerator is an automated system in which the plates containing the milk to be analyzed are placed for incubation. The Accelerator automatically detects the end of the incubation and reads the results. A sample containing antibiotic will be noted as "positive." A sample without antibiotics or with antibiotics at concentrations below detection level will be noted as "negative." The present report includes all technical details about the Delvotest SP NT DA, and the results of the validation study. The validation study demonstrates that the Delvotest SP NT DA conforms to the product performance claims and confirms the robustness of the test. The Delvotest SP NT DA is, therefore, granted Performance Tested Method certification.     

Validation study of the VitaKit A test kit for the determination of vitamin A in fluid milk (2% fat) for routine quality control. Performance Tested Method 061001

A validation study of VitaKit A for quantitation of total vitamin A in 2% fluid milk was carried out according to the guidelines provided by AOAC INTERNATIONAL. The VitaKit A was compared, in terms of repeatability and accuracy, with the U.S. Food and Drug Administration-Interstate Milk Shippers HPLC reference method for determination of total vitamin A in fluid milk with 2% fat. The data obtained by the VitaKit A method are in excellent agreement with the data obtained by the HPLC reference method. Further, a low LOD (0.33 international unit/mL) was obtained for the VitaKit A method; the presence of interferents, like cholesterol and vitamin D3, in the milk had minor influence on the quantitation of total vitamin A by the VitaKit A method. The VitaKit A test kit was found to be stable for 1 year from the date of manufacture when stored at 2-8 degrees C. The method requires 2 h processing time, compared to 1-2 days for the HPLC reference method. The results of this validation study clearly demonstrate that the VitaKit A method is reliable, rapid, and accurate for the quantitation of total vitamin A in fluid milk containing 2% milk fat. An independent study by Q Laboratories Inc., Cincinnati, OH, under the validation guidelines of AOAC INTERNATIONAL, confirmed these findings.     

Reveal Salmonella 2.0 test for detection of Salmonella spp. in foods and environmental samples. Performance Tested Method 960801

Reveal Salmonella 2.0 is an improved version of the original Reveal Salmonella lateral flow immunoassay and is applicable to the detection of Salmonella enterica serogroups A-E in a variety of food and environmental samples. A Performance Tested Method validation study was conducted to compare performance of the Reveal 2.0 method with that of the U.S. Department of Agriculture-Food Safety and Inspection Service or U.S. Food and Drug Administration/Bacteriological Analytical Manual reference culture methods for detection of Salmonella spp. in chicken carcass rinse, raw ground turkey, raw ground beef, hot dogs, raw shrimp, a ready-to-eat meal product, dry pet food, ice cream, spinach, cantaloupe, peanut butter, stainless steel surface, and sprout irrigation water. In a total of 17 trials performed internally and four trials performed in an independent laboratory, there were no statistically significant differences in performance of the Reveal 2.0 and reference culture procedures as determined by Chi-square analysis, with the exception of one trial with stainless steel surface and one trial with sprout irrigation water where there were significantly more positive results by the Reveal 2.0 method. Considering all data generated in testing food samples using enrichment procedures specifically designed for the Reveal method, overall sensitivity of the Reveal method relative to the reference culture methods was 99%. In testing environmental samples, sensitivity of the Reveal method relative to the reference culture method was 164%. For select foods, use of the Reveal test in conjunction with reference method enrichment resulted in overall sensitivity of 92%. There were no unconfirmed positive results on uninoculated control samples in any trials for specificity of 100%. In inclusivity testing, 102 different Salmonella serovars belonging to serogroups A-E were tested and 99 were consistently positive in the Reveal test. In exclusivity testing of 33 strains of non-salmonellae representing 14 genera, 32 were negative when tested with Reveal following nonselective enrichment, and the remaining strain was found to be substantially inhibited by the enrichment media used with the Reveal method. Results of ruggedness testing showed that the Reveal test produces accurate results even with substantial deviation in sample volume or device development time.     

Evaluation of applied biosystems MicroSeq real-time PCR system for detection of Listeria monocytogenes in food. Performance Tested Method 011002

Increasingly, more food companies are relying on molecular methods, such as PCR, for pathogen detection due to their improved simplicity, sensitivity, and rapid time to results. This report describes the validation of a new Real-Time PCR method to detect Listeria monocytogenes in nine different food matrixes. The complete system consists of the MicroSEQ L. monocytogenes Detection Kit, sample preparation, the Applied Biosystems 7500 Fast Real-Time PCR instrument, and RapidFinder Express software. Two sample preparation methods were validated: the PrepSEQ Nucleic Acid extraction kit and the PrepSEQ Rapid Spin sample preparation kit. The test method was compared to the ISO 11290-1 reference method using an unpaired-study design to detect L. monocytogenes in roast beef, cured bacon, lox (smoked salmon), lettuce, whole cow's milk, dry infant formula, ice cream, salad dressing, and mayonnaise. The MicroSEQ L. monocytogenes Detection Kit and the ISO 11290-1 reference method showed equivalent detection based on Chi-square analysis for all food matrixes when the samples were prepared using either of the two sample preparation methods. An independent validation confirmed these findings on smoked salmon and whole cow's milk. The MicroSEQ kit detected all 50 L. monocytogenes strains tested, and none of the 30 nontargeted bacteria strains.     

Performance Tested Method multiple laboratory validation study of ELISA-based assays for the detection of peanuts in food

Performance Tested Method multiple laboratory validations for the detection of peanut protein in 4 different food matrixes were conducted under the auspices of the AOAC Research Institute. In this blind study, 3 commercially available ELISA test kits were validated: Neogen Veratox for Peanut, R-Biopharm RIDASCREEN FAST Peanut, and Tepnel BioKits for Peanut Assay. The food matrixes used were breakfast cereal, cookies, ice cream, and milk chocolate spiked at 0 and 5 ppm peanut. Analyses of the samples were conducted by laboratories representing industry and international and U.S governmental agencies. All 3 commercial test kits successfully identified spiked and peanut-free samples. The validation study required 60 analyses on test samples at the target level 5 microg peanut/g food and 60 analyses at a peanut-free level, which was designed to ensure that the lower 95% confidence limit for the sensitivity and specificity would not be <90%. The probability that a test sample contains an allergen given a prevalence rate of 5% and a positive test result using a single test kit analysis with 95% sensitivity and 95% specificity, which was demonstrated for these test kits, would be 50%. When 2 test kits are run simultaneously on all samples, the probability becomes 95%. It is therefore recommended that all field samples be analyzed with at least 2 of the validated kits.     

Method and kit for adsorbent performance evaluation

Commercial gas separation plants running adsorption processes, including pressure swing adsorption, vacuum pressure swing adsorption and temperature swing adsorption are intended to operate continuously and meet design performance levels over the complete service life of the facility. Performance degradation of the adsorbent materials in extended commercial usage is a common problem. Issues such as adsorbent aging and poisoning by unwanted or unexpected contaminants, represent some of the causes of declining adsorbent performance. Lower adsorption capacity can result in declining productivity and/or product purity for the gas separation plant. Adsorbent troubleshooting is usually accomplished by taking samples from the plant and sending them to off-site laboratories for analyses such as moisture content determination by Karl Fischer titration, or BET surface area or other adsorption capacity measurements. The turnaround time for these laboratory analyses is on the order of days. To short-cut this lengthy process of analysis, we have developed a simple test method and kit for rapid diagnosis of adsorbent performance issues which can be used directly at a plant site. The test method involves the determination of the gas capacity of an adsorbent by equilibrating the adsorbent with an appropriate test gas, deactivating the adsorbent and measuring the amount of test gas released. Once a sample has been acquired, the test can be executed and results obtained in less than 15 min. We show that the test method is accurate to within 5 % of the adsorption capacity determined from isotherm measurements, at equivalent temperature and pressure, and can be used to test common commercial adsorbent types, including low silica zeolites and activated carbons.     

Modification of the BAX System PCR assay for detecting Salmonella in beef, produce, and soy protein isolate. Performance Tested Method 100201

The BAX System PCR assay for Salmonella detection in foods was previously validated as AOAC Research Institute (RI) Performance Tested Method (PTM) 100201. New studies were conducted on beef and produce using the same media and protocol currently approved for the BAX System PCR assay for E. coli O157:H7 multiplex (MP). Additionally, soy protein isolate was tested for matrix extension using the U.S. Food and Drug Administration-Bacteriological Analytical Manual (FDA-BAM) enrichment protocols. The studies compared the BAX System method to the U.S. Department of Agriculture culture method for detecting Salmonella in beef and the FDA-BAM culture method for detecting Salmonella in produce and soy protein isolate. Method comparison studies on low-level inoculates showed that the BAX System assay for Salmonella performed as well as or better than the reference method for detecting Salmonella in beef and produce in 8-24 h enrichment when the BAX System E. coli O157:H7 MP media was used, and soy protein isolate in 20 h enrichment with lactose broth followed by 3 h regrowth in brain heart infusion broth. An inclusivity panel of 104 Salmonella strains with diverse serotypes was tested by the BAX System using the proprietary BAX System media and returned all positive results. Ruggedness factors involved in the enrichment phase were also evaluated by testing outside the specified parameters, and none of the factors examined affected the performance of the assay.     

Validation of the charm 3 SL3 beta-lactam test for screening raw milk in compliance with the U.S. pasteurized milk ordinance. Performance Tested Method 071002

The Charm 3 SL3 beta-Lactam Test is a 3 min receptor-based lateral-flow Rapid One-Step Assay (ROSA) that detects the six beta-lactam drugs of concern approved for dairy cattle in the United States. The method is a biochemical formulation change of the SL3 beta-Lactam Test evaluated and approved in 2007. The Charm 3 SL3 was evaluated under the AOAC Research Institute Performance Tested Method (PTM) program following the protocol of the U.S. Food and Drug Administration, Center for Veterinary Medicine. The method was approved as PTM 071002 on May 8, 2009. The following drugs were detected in three combined lots: penicillin G at 3.8 ppb, ampicillin at 8.0 ppb, amoxicillin at 8.4 ppb, cephapirin at 20.0 ppb, ceftiofur (total metabolites) at 79 ppb, and cloxacillin at 8.6 ppb > or = 90% of the time with 95% confidence. These detection levels are lower than, but within 75% of, the U.S. Safe Level/Tolerances. Lot-to-lot repeatability was typically within 20% of these determined levels. The test kit was found to be suitable for testing thawed frozen samples. It was also found to respond with equal or better sensitivity to samples that contained incurred analytes, i.e., both the microbiologically active parent drug and its active metabolites. There were no interferences from somatic cells at 1.1 million/mL, bacterial cells at 300 000 CFU/mL, or 32 other non-beta-lactam drugs at 100 ppb. Ruggedness experiments indicated that the test procedure is robust. These results meet the fit-for-purpose approval criteria for inclusion in the National Conference for Interstate Milk Shipments milk testing program.     

高效液相色谱法测定动物组织样品中黄曲霉毒素的残留量

采用免疫亲合技术,对动物肝脏和肌肉中黄曲霉毒素残留进行了提取和纯化处理,建立了一种快速、灵敏、简便的测定动物组织中黄曲霉毒素B1、B2、G1、G2的高效液相色谱方法。用反相HPLC分离,柱后光化学衍生,荧光检测器测定黄曲霉毒素含量,保留时间定性,峰面积定量,测定了样品和标准样。结果表明,4种毒素可在10min内完成分离;线性范围为15～1500pg;线性回归系数大于0．9998。用本方法对动物肝脏和肌肉样品进行了加标回收实验,4种黄曲霉毒素的平均回收率为68．71％～83．42％;相对标准偏差为3．51％～7．40％;检出限均达到2．65pg。     

Aflatoxin analogues as possible anticoagulants. III

The synthesis of a series of derivatives of 4-hydroxycoumarin analogues of aflatoxin and a preliminary testing of their anticoagulant activity is described.     

密度泛函理论研究黄曲霉素B1的表面增强拉曼散射效应

用密度泛函理论B3LYP方法和6—311G（d,p）／Lan12DZ优化得到黄曲霉素B1（AFBI）分子及其复合物AFB1-Ag的稳定结构,并计算了复合物的表面增强拉曼光谱和预共振拉曼光谱．结果表明,AFB1分子的拉曼光谱很大程度依赖于吸附位点以及入射光的激发波长．与分子的常规拉曼光谱相比,复合物表面增强拉曼光谱中C=O伸缩振动模的增强因子约为10^2—10^3,是由于复合物的极化率增强而导致的静态化学增强,并分析了振动模式的振动方向与其拉曼强度的关系．选择复合物最大吸收峰附近激发光266和482nm以及远离共振吸收波长785和1064nm作为入射光,计算得到不同入射光激发下复合物的预共振拉曼光谱．结果表明其增强因子最大达N100量级,主要是由电荷转移产生的共振增强引起的．     

Standard Method Performance Requirements

AOAC INTERNATIONAL has developed a process to set Standard Method Performance Requirements (SMPRs). The fitness for intended use of the result of the measurement determines the criteria for these requirements. These criteria are specified by the stakeholder panels made up of key experts from global government, industry, academia, and contract research organizations. This process has been implemented for assays aiming at the detection of bio-threat agents, endocrine disruptors, and quantifying nutrients in infant formula. The process begins with the formation of a balanced stakeholder panel. The stakeholder panel specifies analytes, matrices, current technologies, analytical challenges, and regulatory information at the first meeting. Working groups are formed to provide specific technical expertise to the stakeholder panels. These groups perform the task of translating the specified needs of the stakeholder panel into concise measurement requirements. The draft SMPRs go back to the stakeholder panel for review and approval and then are posted for comment on the AOAC web site. These requirements become consensus requirements that have been developed with openness, balance of interests, an appeals procedure, consensus, and transparency.     

HPLC与ELISA法测定牛奶中黄曲霉毒素M1的比较研究

对HPLC法和ELISA法检测牛奶样品中黄曲霉毒素M1(AFM1)残留量进行了比较,探讨了牛奶中黄曲霉毒素M1的最佳检测方法。结果显示,对于已知加标样品(阳性),HPLC法回收率为68%～85%;ELISA法回收率为97%～109%,HPLC法和ELISA法具有良好的相关性。ELISA法检验成本较低、检测迅速,但可能出现少量的假阳性和假阴性,故可用于待测样品的初筛;HPLC法检测结果准确,但检测费用较高,适用于黄曲霉毒素M1残留的确证。     

UPLC-MS-MS法快速测定玉米中的黄曲霉毒素B1

采用高效液相色谱-串联质谱法检测玉米中的黄曲霉毒素B1。对样品前处理条件、净化和分离条件进行了优化,黄曲霉毒素B1检出限为0．02μg／kg,回收率为83．8％～95．7％。建立的方法简便、快速、灵敏度高,能够满足玉米中痕量黄曲霉毒素B1农残的高灵敏测定要求。     

Roche/BIOTECON Diagnostics LightCycler foodproof L. monocytogenes detection kit in combination with ShortPrep foodproof II Kit. Performance-Tested Method 070401

A method was developed for the detection of L. monocytogenes in food based on real-time polymerase chain reaction (PCR). This advanced PCR method was designed to reduce the time needed to achieve results from PCR reactions and to enable the user to monitor the amplification of the PCR product simultaneously, in real-time. After DNA isolation using the Roche/BIOTECON Diagnostics ShortPrep foodproof II Kit (formerly called Listeria ShortPrep Kit) designed for the rapid preparation of L. monocytogenes DNA for direct use in PCR, the real-time detection of L. monocytogenes DNA is performed by using the Roche/BIOTECON Diagnostics LightCycler foodproof L. monocytogenes Detection Kit. This kit provides primers and hybridization probes for sequence-specific detection, convenient premixed reagents, and different controls for reliable interpretation of results. For repeatability studies, 20 different foods, covering the 15 food groups recommended from the AOAC Research Institute (AOAC RI) for L. monocytogenes detection were analyzed: raw meats, fresh produce/vegetables, processed meats, seafood, egg and egg products, dairy (cultured/noncultured), spices, dry foods, fruit/juices, uncooked pasta, nuts, confectionery, pet food, food dyes and colorings, and miscellaneous. From each food 20, samples were inoculated with a low level (1-10 colony-forming units (CFU)/25 g) and 20 samples with a high level (10-50 CFU/25 g) of L. monocytogenes. Additionally, 5 uninoculated samples were prepared from each food. The food samples were examined with the test kits and in correlation with the cultural methods according to U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) or U.S. Department of Agriculture (USDA)/Food Safety and Inspection Service (FSIS) Microbiology Laboratory Guidebook. After 48 h of incubation, the PCR method in all cases showed equal or better results than the reference cultural FDA/BAM or USDA/FSIS methods. Fifteen out of 20 tested food types gave exactly the same amount of positive samples for both methods in both inoculation levels. For 5 out of 20 foodstuffs, the PCR method resulted in more positives than the reference method after 48 h of incubation. Following AOAC RI definition, these were false positives because they were not confirmed by the reference method (false-positive rate for low inoculated foodstuffs: 5.4%; for high inoculated foodstuffs: 7.1%). Without calculating these unconfirmed positives, the PCR method showed equal sensitivity results compared to the alternative method. With the unconfirmed PCR-positives included into the calculations, the alternative PCR method showed a higher sensitivity than the microbiological methods (low inoculation level: 100 vs 98.0%; sensitivity rate: 1; high inoculation level: 99.7 vs 97.7%; sensitivity rate, 1). All in-house and independently tested uninoculated food samples were negative for L. monocytogenes. The ruggedness testing of both ShortPrep foodproof II Kit and Roche/BIOTECON LightCycler foodproof L. monocytogenes Detection Kit showed no noteworthy influences to any variation of the parameters component concentration, apparatus comparison, tester comparison, and sample volumes. In total, 102 L. monocytogenes isolates (cultures and pure DNA) were tested and detected for the inclusivity study, including all isolates claimed by the AOAC RI. The exclusivity study included 60 non-L. monocytogenes bacteria. None of the tested isolates gave a false-positive result; specificity was 100%. Three different lots were tested in the lot-to-lot study. All 3 lots gave equal results. The stability study was subdivided into 3 parts: long-term study, stress test, and freeze-defrost test. Three lots were tested in 4 time intervals within a period of 13 months. They all gave comparable results for all test intervals. For the stress test, LightCycler L. monocytogenes detection mixes were stored at different temperatures and tested at different time points during 1 month. Stable results were produced at all storage temperatures. The freeze-defrost analysis showed no noteworthy aggravation of test results. The independent validation study examined by Campden and Chorleywood Food Research Association Group (CCFRA) demonstrated again that the LightCycler L. monocytogenes detection system shows a comparable sensitivity to reference methods. With both the LightCycler PCR and BAM methods, 19 out of 20 inoculated food samples were detected. The 24 h PCR results generated by the LightCycler system corresponded directly with the FDA/BAM culture results. However, the 48 h PCR results did not relate exactly to the FDA/BAM results, as one sample found to be positive by the 48 h PCR could not be culturally confirmed and another sample which was negative by the 48 h PCR was culturally positive.     

Performance of an ultra-low elution-volume 96-well plate: drug discovery and development applications

Recently, sample preparation has been considered to be the major cause of bottlenecks during high-throughput analysis. With the assistance of robotic liquid handlers and the 96-well plate format, more samples can be prepared for subsequent liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis. Protein precipitation is still widely used despite potential loss of sensitivity or variable results due to ion suppression. The use of solid-phase extraction (SPE) clearly gives superior results but may not be as cost effective as protein precipitation due to the labor and material costs associated with the process. Here, a novel 96-well SPE plate is described that was designed to minimize the elution volume required for quantitative elution of analytes. The plate is packed with 2 mg of a high-capacity SPE sorbent that allows loading of up to 750 microL of plasma, while the novel design permits elution with as little as 25 microL. Therefore, the plate offers up to a 15-fold increase in sample concentration. The evaporation and reconstitution step that is typically required in SPE is avoided due to the concentrating ability of the plate. Examples of applications in drug discovery/development are shown and results are compared to protein precipitation. Excellent sensitivity and linearity are demonstrated.     

Modification of the BAX Salmonella test kit to include a hot start functionality (modification of AOAC Official Method 2003.09)

In 2010, the BAX System PCR assay for Salmonella was modified to include a hot start functionality designed to keep the reaction enzyme inactive until PCR begins. To validate the assay's Official Methods of Analysis status to include this procedure modification, an evaluation was conducted on four food types that were simultaneously analyzed with the BAX System and either the U.S. Food and Drug Administration's Bacteriological Analytical Manual or the U.S. Department of Agriculture-Food Safety and Inspection Service Microbiology Laboratory Guidebook reference method for detecting Salmonella. Identical performance between the BAX System method and the reference methods was observed. Additionally, lysates were analyzed using both the BAX System Classic and BAX System Q7 instruments with identical results using both platforms for all samples tested. Of the 100 samples analyzed, 34 samples were positive for both the BAX System and reference methods, and 66 samples were negative by both the BAX System and reference methods, demonstrating 100% correlation. No instrument platform variation was observed. Additional inclusivity and exclusivity testing using the modified test kit demonstrated the test kit to be 100% accurate in evaluation of test panels of 352 Salmonella strains and 46 non-Salmonella strains.     

高效液相色谱法测定阿卡波糖水解产物中的Valienamine含量

1引言Valienam ine((1S,2S,3S,4R)-1-氨基-5-(羟甲基)环己-5-烯-2,3,4-三元醇),分子式C7H13NO4,是一种由假单孢菌降解井冈霉素获得的假氨基糖,可通过阿卡波糖或井冈霉素水解来制备。它不仅能够有效治疗糖尿病及高血糖所导致的肥胖症,而且还能治疗艾滋病等其它疾病,所以对其的研究和开发已引起全世界生物医学领域的广泛关注。Valienam ine的检测方法主要有糖化酶检测法、紫外分光光度法、质谱法和薄层扫描法等,这些方法都存在难于定量或定量不准确、检测不方便的缺陷。有人用p-硝基氟苯与Valienam ine反应衍生生成N-p-硝基苯-Valienam …     

低温燃烧法制备Ce-TiO2光催化材料与性能研究

以硫酸钛为原料,采用低温燃烧法制备Ce掺杂TiO2光催化材料,借助SEM,XRD对样品进行表征。以粉体晶粒粒径和晶型组成为指标确定Ce最佳掺杂浓度为0.3%(摩尔分数)。选取10 mg.L-1的罗丹明B溶液,粉体添加量为1 g.L-1,高压汞灯照射下光催化降解3 h,罗丹明B的降解率为49.15%。选取光照时间1 h,粉体添加量10 mg,菌液浓度103 cfu.ml-1,普通日光灯为光源,检测产品的杀菌率为34.90%,Ce-TiO2粉体的催化活性明显优于纯TiO2。     

芝麻中芝麻素含量的高效液相色谱法测定

建立了一种快速、简便测定芝麻中芝麻素含量的方法。将芝麻粉碎后用甲醇超声提取,优化了粉碎粒径、超声料液比和提取时间。结果表明,芝麻样品经粉碎及超声萃取后,采用高效液相色谱法(HPLC)测定,色谱柱为C18(4.6 mm×250 mm,5μm),流动相为甲醇-水(75∶25),流速1.0 mL/min,柱温30℃,检测波长286 nm。芝麻素在0.043 84~0.569 9μg范围内线性关系良好,回归方程为A=1 346 015m+2 136,r2=0.999 9;芝麻素的加标回收率为100%,RSD为1.1%,不同产地芝麻中的芝麻素含量测定结果为0.260%~0.502%。该方法具有操作简便、稳定、专属、可重复的特点,可用于芝麻的质量控制。