不同品牌霉菌毒素检测试剂盒的质量评价

目的评价当前国内外应用较广的9个不同品牌黄曲霉毒素B1、玉米赤霉烯酮、呕吐毒素检测试剂盒的准确性。方法选取3种不同浓度的质控样品、4种饲料原料样品(玉米干全酒精糟、玉米胚芽粕、麦麸、豆粕)、2种饲料产品样品(蛋鸡配合饲料、牛浓缩饲料)。采用3种不同浓度的质控样(用萃取液点板6个孔)的检测结果变异系数(variable coefficient, CV)作为精密度评价指标。采用质控样的6个单独检测结果变异系数作为系统的精密度评价指标,采用实际样品的检测结果评价试剂盒的适用性。结果呕吐毒素试剂盒CV值平均值2号品牌2.7%, 3号品牌3.4%。玉米赤霉烯酮试剂盒CV值平均值3号品牌4.6%, 2号品牌5.5%, 4号品牌5.4%。黄曲霉毒素B1试剂盒CV值平均值5号品牌3.3%, 8号品牌3.7%, 2号品牌4.1%。结论依据试剂盒精密度、准确度、灵敏度和适应性综合评估2号品牌呕吐毒素试剂盒综合结果优秀, 9号品牌玉米赤霉烯酮试剂盒综合结果较好, 5号品牌黄曲霉毒素B1综合结果较好。同时,就检测上述3种常见毒素而言,没有发现一个品牌的试剂盒可以同时在多个毒素检测上表现优秀。     

酶联免疫法测定食品和饲料中的黄曲霉毒素

本实验应用酶联免疫方法对食品和饲料中的黄曲霉毒素进行测定,奶粉中黄曲霉毒素M1的多次重复测定的结果的变异系数小于12%,回收率接近90%。说明本试剂盒测定结果比较精确,重现性较好。食品辅料及饲料中的黄曲霉毒素B1的测定结果的变异系数小于15%,说明实验结果的重现性较好。     

Inter-laboratory validation study of five commercial ELISA test kits for the determination of peanut proteins in biscuits and dark chocolate.

The results of an inter-laboratory study with five commercially available peanut ELISA test kits to detect and quantify peanut residues in two food matrices (biscuit and dark chocolate) at four different concentrations (0-10 mg peanut kg(-1) matrix corresponding to about 0-2.5 mg peanut protein kg(-1) matrix) are reported. In general the five ELISA test kits evaluated could detect peanut protein in the two food matrices. In three cases, the study challenged the test kits beyond their intended use for quantification below the manufacturers' defined cut-off limits. Generally, all five ELISA test kits performed well in the concentration range 5-10 mg kg(-1) rather than in the low concentration range (2.0 or 2.5 mg kg(-1)). The variation in the found recoveries of peanut between the different test kits had a spread of 44-191% across all concentrations. The quantification characteristics between test kits differed significantly at the very low mg kg(-1) level. Two test kits performed well even at concentrations below 5 mg kg(-1) with reproducibilities of 27-36% for biscuits and 45-57% for chocolate.     

Measuring concentrations of insulin-like growth factor-I with an enzyme-linked immunosorbent antibody assay in plasma samples from Holstein cows

This study describes the use of a commercial nonextraction ELISA to quantify total concentrations of insulin-like growth factor-I (IGF-I) in plasma samples from Holstein cows fed pasture-based diets varying in dry matter and metabolizable energy density. The assays were conducted using the protocols provided by the manufacturer. The ELISA was verified for linearity, accuracy in measuring IGF-I from spiked plasma samples, and precision involving variation within and between assays. Validation also involved comparing results of the ELISA against an established RIA after defatted plasma samples were subjected to acid gel HPLC to dissociate and separate IGF and IGF binding protein complexes. Validation results had low coefficients of variation (CV; intraassay CV of less than 6% and interassay CV of less than 8%) and a high recovery percentage of IGF-I (79%) from samples spiked with unbound IGF-I. The coefficients of determination between the ELISA and the RIA reference assay were 0.90 in 2 separate assays. Associations between the RIA and the ELISA were higher, and the limits of agreement at 95% confidence intervals were narrower compared with those between the RIA and a modified ELISA procedure in which IGF binding protein complexes were extracted using HPLC. The samples were obtained from cows sampled over a period of 5 wk. These results demonstrated that the DSL-10-2800 nonextraction IGF-I ELISA was acceptably specific and sufficiently sensitive to be used to measure the extent and patterns of change in the plasma concentrations of IGF-I in samples from lactating Holstein cows.     

Whipping test for the determination of foaming capacity of protein: a collaborative study

A standard whipping test, which involved preparing a 0.5% protein dispersion, whipping the dispersion in a Kenwood Chef food mixer and measuring the foaming properties (foam expansion, FE; foam volume stability, FVS; foam liquid stability, FLS) was collaboratively evaluated. A total of sixty determinations was made involving six laboratories, each of which analysed five samples in duplicate. The repeatability coefficient of variation ( CV ₀) in the estimation of FE and FVS properties of proteins was 6.25%. The reproducibility coefficient of variation ( CV _X) for FE and FVS was 9.8%, excluding the FE value (i.e. 20%) for spray-dried albumen. The CV ₀ and CV _X values for FLS properties of proteins ranged from 7.4 to 13.4% and 17.2 to 24.6%, respectively. Thus, FE, FVS and FLS properties can be used for intralaboratory comparisons (e.g. to determine batch-to-batch variation in commercial protein products), whereas FE and FVS values are more reliable for inter-laboratory comparisons of the foaming properties of proteins.     

Dual-energy X-ray absorptiometry of the pig: Protocol development and evaluation

As part of a prospective study in bone mineralisation in adult pigs it was necessary to establish guidelines and to define sites for bone mineral measurements. Particular requirements were that, the protocol should be suitable for a mass screening programme in both postmortem specimens and in live animals, and should deliver results of known reliability. Estimates of bone mineral content (BMC) and bone mineral density (BMD) in areas within the 4th metacarpal bone yielded coefficients of variation (CV) in the order of 7% for both regions and estimates in regions which included the entire metacarpal-phalangeal area yielded CV values in the order of 0.7% and 0.6% for BMC and BMD, respectively. A region of interest taken from the coccygeal vertebrae yielded coefficient of variation values of 3% and 2% for BMC and BMD, respectively. Accuracy of dual-energy X-ray absorptiometry (DXA) was estimated using a standard curve derived from BMC determined by ashing. There was a high correlation between mineral content determined by DXA and by ashing (R(2)=0.99, p<0.0001). The results suggest that the regions used in this study are suitable for use in large, mass screening, prospective studies.     

Evaluation of two new arsenic field test kits capable of detecting arsenic water concentrations close to 10 microg/L

Millions of people worldwide are exposed to arsenic-contaminated drinking water. Arsenic field test kits may offer a cost-effective approach for measuring these exposures in the field, although the accuracy of some kits used in the past has been poor. In this study, arsenic concentrations were measured in 136 water sources in western Nevada using two relatively new arsenic test kits and compared to laboratory measurements using atomic fluorescence spectroscopy (AFS). Spearman's rank correlation coefficients comparing the Quick Arsenic and Hach EZ kits to laboratory measurements were 0.96 (p < 0.001) and 0.95 (p < 0.001), respectively. When analyzed in seven exposure categories (0-9, 10-19, 20-49, 50-99, 100-199, 200-499, and > or = 500 microg/L), test kit and AFS measurements were in the same category in 71% (Quick Arsenic) and 62% (Hach EZ) of samples, and within one category of each other in 99% (Quick Arsenic) and 97% (Hach EZ) of samples. Both kits identified all water samples with high arsenic concentrations (> 15 microg/L) as being above the United States Environmental Protection Agency's drinking water standard and the World Health Organization's guideline value for arsenic of 10 microg/L. These results suggestthatthese easily portable kits can be used to identify water sources with high arsenic concentrations and may provide an important tool for arsenic surveillance and remediation programs.     

Collaborative test of the fluorimetric determination of selenium in a test solution, milk powder and bovine liver

Results are reported for an inter-laboratory test conducted to assess precision (repeatability, reproducibility) and accuracy of a collaborative method for the fluorimetric determination of selenium (Se). The seven participating laboratories analysed one test solution, four samples of milk powder and two samples of freeze-dried bovine liver. Each set of samples comprised three duplicates: two colorant-disguised milk powders and one code disguised freeze-dried bovine liver. Two of the milk powders were enriched with 90.7 micrograms/kg Se as seleno-DL-methionine. One set of results had to be rejected because the laboratory involved did not adhere to the collaborative method. Results from a second laboratory contained both stragglers and outliers. The five remaining laboratories performed the method satisfactory. Results from these laboratories were statistically evaluated according to ISO 5725. The average coefficient of variation within a laboratory (repeatability) was 4.8% and between laboratories (reproducibility) 6.0% for the milk powder and bovine liver samples. Recovery for the test solution, target value 120 micrograms Se/1, was 96% and the average recovery for the Se enriched milk powder was 88%. The mean result for the milk powder was 98.9 micrograms/kg (n = 10), coefficient of variation (CV) 6.7%, and for Se enriched milk powder 178.3 micrograms/kg, coefficient of variation (CV) 3.6%. For freeze-dried bovine liver, these results were 238.4 micrograms/kg and 4.1% respectively.     

3种β-兴奋剂化学发光酶免疫试剂盒分析检测方法的评价

选取3种市售β-兴奋剂化学发光酶免疫分析检测试剂盒,从线性关系、灵敏度、选择性、检出限、准确度、重复性及变异性等方面进行实验,并经农业部1025号公告—18—2008中高效液相色谱—串联质谱法验证比较,初步建立了一套化学发光酶免疫分析检测试剂盒质量评价方法。结果表明:沙丁胺醇、盐酸克伦特罗、莱克多巴胺3种试剂盒在测定范围内线性关系良好,灵敏度分别为0.7、1.8、1.0 ng/mL,检出限分别为0.2、0.1、0.2 ng/mL,猪肝、猪肉和牛肉的回收率在81.9%~115.2%之间,重复性变异系数为3.3%~9.7%,批间变异系数为9.9%~16.5%,交叉反应率均符合产品说明书要求,可以用于猪肝、猪肉、牛肉样品中沙丁胺醇、盐酸克伦特罗、莱克多巴胺的残留检测。     

呋喃西林代谢物单克隆抗体及荧光免疫层析试纸条的制备

为建立一种基于荧光免疫层析技术的呋喃西林代谢物的快速检测方法,采用活性酯法合成呋喃西林代谢物（氨基脲盐酸盐）（semicarbazide hydrochloride,SEM）人工抗原,并获得SEM的单克隆抗体,通过将量子点微球（quantumdot submicrobeads,QBs）与SEM偶联,得到QBs探针,制备荧光免疫层析试纸条。结果表明：SEM免疫荧光层析试纸条检测鱼肉组织的检测限为0.247 μg/L,不同添加质量浓度的回收率均在70%～120%之间,批内、批间变异系数均在15%以下,符合我国动物源食品中兽药残留检测方法相关指导文件的规定,且在2～8 ℃条件下可以保存6 个月以上,稳定性良好。     

Compositional differences in some limabean (Phaseolus lunatus) varieties

Eighteen varieties of limabean (Phaseolus lunatus) were analysed for their proximate chemical composition, mineral constituents and amino acid composition. The crude protein levels were remarkably similar except for that of TPL 071-33 (25·9%), which was higher than for most of the other varieties. Ether extract was low in all varieties, with a percentage coefficient of variation of 46·2%. Potassium was the most abundant mineral element, ranging from 550 mg/100 g in TPL 4 to 2380 mg/100 g in TPL 14e. Phosphorus and magnesium contents were also appreciably high in all varieties while calcium was the poorest major mineral element. Methionine and cystine were the most limiting amino acids and showed some degree of intervarietal differences, their coefficients of variation (CV) being 19·0 and 12·2%, respectively. Other amino acids which showed varietal differences included serine (CV 18·2%), glutamic acid (CV 15·4%), proline (CV 14·4%), glycine (CV 14·5%) and alanine (CV 14·4%). Limabean TPL 2 was deficient in histidine whilst trytophan was adequate in all varieties but deficient in TPL 2, TPL 6, TPL 13 and TPL 187. Glutamic acid, followed by aspartic acid, levels were generally the highest in each of the limabean varieties. The implications of these findings are fully discussed.     

Evaluation of three commercial ELISA kits for rapid screening of deoxynivalenol in unpolished wheat

Evaluation of commercial ELISA kits for the screening of deoxynivalenol (DON) was carried out. Three kinds of commercial kits supplied by different companies were used. Three lots of naturally contaminated wheat and DON-free wheat (<0.05 microg/g) were used as samples. The values obtained from ELISA were compared with those of the HPLC-UV method. In the results obtained from 14 institutions using ELISA, the CV values were less than 17.6% for all the commercial kits. The DON-free sample was not detected as positive with any commercial kits. Also there was no negative finding among the naturally contaminated samples used in this experiment. Coefficients of determination in ELISA and HPLC analysis were 0.979-0.999. These results indicate that ELISA using any of the three commercial kits is efficient for the screening of DON in wheat.     

TENSIPRESSER PRECISION IN MEASURING COOKED RICE ADHESIVENESS

Precision in measuring cooked rice adhesiveness was studied using a Tensipresser and the "full cup" method. To determine the optimum adhesiveness parameter, 14 samples each from rice harvests in 1992 and 1993 were used. Out of the 42 parameters derived from a force-distance curve, the ratio of cumulative adhesiveness to cumulative hardness,     was used as the optimum because of its high correlation (r >0.9) with sensory adhesiveness, high precision (mean coefficient of variation (CV) of 14 samples <15% and maximum CV of 14 samples <25 %), and high resolution based on Kruskal-Wallis test statistic H. For parameter     , the maximum CV decreased with the increasing number of bites, and statistic H indicated that increasing the number of bites from 1 to 5 increased resolution, but bites exceeding 5 did not clearly do so.     

Skatole and indole concentrations in Longissimus dorsi and fat samples of pigs

A HPLC-NP (normal phase-high performance liquid chromatography) method for determining the concentration of skatole and indole in Longissimus dorsi samples is described. Lipids containing skatole and indole were extracted in chloroform:methanol (2:1) at room temperature and dehydrated by liquid-liquid extraction with an aqueous solution saturated with 10% of sodium chloride. The organic phase was evaporated to dryness and redissolved in 10 ml of hexane:2-propanol (92:8). Indolic compounds were separated on a Hypersil aminopropylsilica column (5 μm) (250×4.6 mm i.d.). The mobile phase was hexane:2-propanol (92:8) and detection was by fluorescence (excitation at 280 nm and emission at 360 nm). Linearity was found in the range of 0.05-0.4 μg/g and the coefficient of correlation was above 0.99 for both compounds. The within day (n=5) variation was at 0.05, 0.2 and 0.4 μg/g and the CV (coefficient of variation) values for relative areas determined at these concentrations were less than 13%. This method was used to compare the concentrations of skatole and indole in different samples: L. dorsi muscle, the fat covering the L. dorsi and subcutaneous fat. A correlation was observed between the concentration of indole and skatole in the back fat and fat covering the L. dorsi samples (P<0.001, r=0.99). No significant correlation was obtained in L. dorsi samples, between skatole and indole levels. In spite of the correlation shown between skatole and indole concentrations in the back fat and L. dorsi samples, the mean concentrations of these compounds were significantly higher (P<0.05) in the back fat samples.     

Effects on milk yield of milking interval regularity and teat cup attachment failures with robotic milking systems

A database consisting of 35291 milking records from 83 cows was built over a period of 10 months with the objectives of studying the effect of teat cup attachment failures and milking interval regularity on milk production with an automated milking system (AMS). The database collected records of lactation number, days in milk (DIM), milk production, interval between milkings (for both the entire udder and individual quarters in case of a teat cup attachment failure) and average and peak milk flows for each milking. The weekly coefficient of variation (CV) of milking intervals was used as a measure of milking regularity. DIM, milking intervals, and CV of milking intervals were divided into four categories coinciding with the four quartiles of their respective distributions. The data were analysed by analysis of variance with cow as a random effect and lactation number, DIM, the occurrence of a milking failure, and the intervals between milkings or the weekly CV of milking intervals as fixed effects. The incidence of attachment failures was 7.6% of total milkings. Milk production by quarters affected by a milking failure following the failure was numerically greater owing to the longer interval between milkings. When accounting for the effect of milking intervals, milk production by affected quarters following a milking failure was 26% lower than with regular milkings. However, the decrease in milk production by quarters affected by milking failures was more severe as DIM increased. Average and peak milk flows by quarters affected by a milking failure were lower than when milkings occurred normally. However, milk production recovered its former level within seven milkings following a milking failure. Uneven frequency (weekly CV of milking intervals >27%) decreased daily milk yield, and affected multiparous more negatively than primiparous cows.     

乳制品中沙门氏菌分子检测方法的验证研究

目的验证沙门氏菌、非沙门菌及阳性核酸模板对MICROFAST?沙门氏菌核酸检测试剂盒(PCR-探针法)的特异性和稳定性,同时比对试剂盒法与GB 4789.4—2016培养法定性检测结果的一致性。方法用实验室保存的30株沙门氏菌和15株非沙门氏菌菌株验证MICROFAST?沙门氏菌核酸检测试剂盒(PCR-探针法)的特异性。通过人工添加不同浓度沙门氏菌对30个乳制品样本,采用国家标准法和试剂盒法同时检验,探究方法的一致性。选择制备好的5份阳性核酸模板,每个模板分别使用3个批次的试剂盒进行检测,对实验结果进行重复性和显著性分析,确定不同试剂盒批次间是否存在显著性差异。结果 30株沙门氏菌菌株和15株非沙门氏菌菌株的特异性检测结果表明,MICROFAST?沙门氏菌核酸检测试剂盒(PCR-探针法)对沙门氏菌的特异性符合预期。人工添加的阳性样本检测结果表明,在乳制品样本范围内,试剂盒的假阳性率与假阴性率为0。3个批次的试剂盒对5份阳性模板检测结果之间没有显著性差异,变异系数CV均小于1%。结论该试剂盒方法与国家标准法相比,具有操作简单、快速等优势,适合乳制品加工过程微生物快速检测。     

Quantification of Free 2-Furfurylthiol in Coffee Brew Using a Prefabricated Coffee Model

A reliable and sensitive method was developed to quantify the free 2-furfurylthiol (2-FFT) in coffee brew. Using cysteine addition combined with vacuum distillation, a coffee model with trace 2-FFT and similar physicochemical properties as coffee brew was prepared for standard curve construction. During the model preparation, 2-FFT in bound form was released to free 2-FFT and removed from coffee brew through vacuum distillation. The method sensitivity, precision, accuracy and selectivity were evaluated. The limit of quantification (LOQ) was 3.0 μg/L. The coefficient of variation (CV) was 7.1%. The average recovery rates were 86.8–106.2% at spiked concentrations of 22.6, 135.8 and 181 μg/L. Coffee brews prepared using Robusta and Arabica roasted coffee beans were analyzed, and results showed that coffee brew from Robusta coffee beans had a greater amount of free 2-FFT (20.94 μg/L) than Arabica coffee beans from Yunnan (11.34 μg/L) and Columbia (15.33 μg/L), respectively. This method would be beneficial to investigate the free amount of 2-FFT in coffee brew and the sensorial assay based on free 2-FFT concentration.     

Effects of feeding rumen-protected methionine pre- and postpartum in multiparous Holstein cows: Lactation performance and plasma amino acid concentrations

Objectives were to evaluate the effect of feeding rumen-protected methionine (RPM) in pre- and postpartum total mix ration (TMR) on lactation performance and plasma AA concentrations in dairy cows. A total of 470 multiparous Holstein cows [235 cows at University of Wisconsin (UW) and 235 cows at Cornell University (CU)] were enrolled approximately 4 wk before parturition, housed in close-up dry cow and replicated lactation pens. Pens were randomly assigned to treatment diets (pre- and postpartum, respectively): UW control (CON) diet = 2.30 and 2.09% of Met as percentage of metabolizable protein (MP) and RPM diet = 2.83 and 2.58% of Met as MP; CU CON = 2.22 and 2.19% of Met as percentage of MP, and CU RPM = 2.85 and 2.65% of Met as percentage of MP. Treatments were evaluated until 112 ± 3 d in milk (DIM). Milk yield was recorded daily. Milk samples were collected at wk 1 and 2 of lactation, and then every other week, and analyzed for milk composition. For lactation pens, dry matter intake (DMI) was recorded daily. Body weight and body condition score were determined from 4 ± 3 DIM and parturition until 39 ± 3 and 49 DIM, respectively. Plasma AA concentrations were evaluated within 3 h after feeding during the periparturient period [d ?7 (±4), 0, 7 (±1), 14 (±1), and 21 (±1); n = 225]. In addition, plasma AA concentrations were evaluated (every 3 h for 24 h) after feeding in cows at 76 ± 8 DIM (n = 16) and within 3 h after feeding in cows at 80 ± 3 DIM (n = 72). The RPM treatment had no effect on DMI (27.9 vs. 28.0 kg/d) or milk yield (48.7 vs. 49.2 kg/d) for RPM and CON, respectively. Cows fed the RPM treatment had increased milk protein concentration (3.07 vs. 2.95%) and yield (1.48 vs. 1.43 kg/d), and milk fat concentration (3.87 vs. 3.77%), although milk fat yield did not differ. Plasma Met concentrations tended to be greater for cows fed RPM at 7 d before parturition (25.9 vs. 22.9 µM), did not differ at parturition (22.0 vs. 20.4 µM), and were increased on d 7 (31.0 vs. 21.2 µM) and remained greater with consistent concentrations until d 21 postpartum (d 14: 30.5 vs. 19.0 µM; d 21: 31.0 vs. 17.8 µM). However, feeding RPM decreased Leu, Val, Asn, and Ser (d 7, 14, and 21) and Tyr (d 14). At a later stage in lactation, plasma Met was increased for RPM cows (34.4 vs. 16.7 µM) consistently throughout the day, with no changes in other AA. Substantial variation was detected for plasma Met concentration (range: RPM = 8.9–63.3 µM; CON = 7.8–28.8 µM) among cows [coefficient of variation (CV) > 28%] and within cow during the day (CV: 10.5–27.1%). In conclusion, feeding RPM increased plasma Met concentration and improved lactation performance via increased milk protein production.     

Associations between routinely collected Dairy Herd Improvement data and insemination outcome in UK dairy herds

Milk constituent concentrations in samples taken during early lactation are often used to generate proxy measures for energy balance in dairy herds. This study aimed to explore associations between these and other measures routinely recorded by Dairy Herd Improvement schemes and insemination outcome, with an emphasis on the likely predictiveness of such measures for conception risk (the proportion of inseminations that are successful) at herd level. Data from 312 UK dairy herds were restructured so that each unit of data represented an insemination at less than 100 d in milk (DIM). Milk constituent concentrations from the first and second test days (corrected for the effects of season and DIM at sampling) were used as potential predictors of insemination outcome in a logistic regression model. Other predictors included representations of milk yield and other information routinely collected by Dairy Herd Improvement Associations; random effects were used to account for clustering at cow and herd levels. The final model included a large number of predictors, with several interaction and nonlinear terms. The relative effect sizes of the measures of early lactation milk constituent concentrations were small. The full model predicted just under 64% of observed variation in herd-year conception risk (i.e., the proportion of inseminations that were successful in each herd in each calendar year); however, around 40% was accounted for by the herd-level random effect. The predictors based on early lactation milk constituent concentrations accounted for less than 0.5% of observed variation, and representations of milk yield (both overall level of yield and early lactation curve shape) accounted for around 7%; DIM at insemination, parity, interservice interval, year, and month accounted for the remaining 15%. These results suggest that early lactation milk constituent information is unlikely to predict herd conception risk to a useful extent. The large proportion of observed variation explained by the herd-level random effect suggests that there are unmeasured (in this study) or unmeasurable factors that are consistent within a herd and are highly influential in determining herd conception risk.     

Inter- and Intra-Laboratory Variation in Amino Acid Analysis of Food Proteins

Amino acid compositions of casein, egg white, beef, soy isolate, rapeseed concentrate, pea flour and wheat flour were studied by seven collaborating laboratories. Samples were hydrolyzed with 6N HCl, performic acid + 6N HCl, and (with one exception) 4.2N NaOH. Amino acids were then determined by ion-exchange chromatography using automatic analyzers. One laboratory, however, determined tryptophan by treatment with p-dimethylaminobenz-aldehyde. The colorimetric determination of tryptophan was comparable to the tryptophan values determined by ion-exchange chromatography. Interlaboratory variation of tryptophan (coefficients of variation, CV, up to 24%), cystine and methionine (CV up to 17%), was greater than that of most other amino acids (CV up to 10%). Intralaboratory variation for all amino acids was less than 5%.