Determination of melamine in milk and dairy products by high performance liquid chromatography

A simple, precise, accurate, and validated reverse-phase HPLC method was developed for the determination of melamine in milk (pasteurized and UHT milk) and dairy products (powdered infant formula, fruit yogurt, soft cheese, and milk powder). Following extraction with acetonitrile:water (50:50, vol/vol), samples were purified by filter (0.45 μm), separated on a Nucleosil C8 column (4.6 mm × 250 mm, 3 μm) with acetonitrile:10 mmol/L sodium L-octane sulfonate (pH 3.1; 15:85, vol/vol) as mobile phase at a flow rate of 1 mL/min, and determined by a photodiode array detector. A linear calibration curve was obtained in the concentration range from 0.05 to 5 mg/kg. Milk and dairy products were fortified with melamine at 4 levels producing average recovery yields of 95 to 109%. The limits of detection and quantification of melamine were 35 to 110 and 105 to 340 μg/kg, respectively. The method was then used to analyze 300 samples of milk and dairy products purchased from major retailers in Turkey. Melamine was not found in infant formulas and pasteurized UHT milk, whereas 2% of cheese, 8% of milk powder, and 44% of yogurt samples contained melamine at the 121, 694±146, and 294±98 μg/kg levels, respectively. These findings were below the limits set by the Codex Alimentarius Commission and European Union legislation. This is the first study to confirm the existence of melamine in milk and dairy products in Turkey. Consumption of foods containing these low levels of melamine does not constitute a health risk for consumers.     

毛细管电泳法对乳及乳制品中真蛋白的测定

利用毛细管电泳法对市售乳清蛋白粉、采自当地奶牛场的牛初乳及原料奶、同一品牌不同阶段或同一阶段内不同厂家、不同国别的婴儿配方奶粉、不同保质期的液态奶、不同国别的超高温灭菌(UHT)奶和酸奶、国产复原乳中α- 乳白蛋白(α-Lac)、β- 乳球蛋白A (β-LgA)及β- 乳球蛋白B (β-LgB)的质量分数进行测定。结果表明：α-Lac、β-LgA 及β-LgB 的质量分数在乳清蛋白粉、牛初乳及原料奶中依次降低。液态奶中上述3 种蛋白的质量分数与保质期有关,一般保质期越久的产品,上述3 种蛋白的质量分数越低。大多数酸奶中α-Lac 的质量分数均高于UHT 奶。婴儿配方奶粉中α-Lac 质量分数随着阶段数的增加而增加,而且同一阶段内不同厂家不同国别配方奶粉中α-Lac 质量分数差异也较大,并非进口奶粉中α-Lac 质量分数均比国产奶粉高。     

Short communication: Aflatoxin M1 in dairy products sold in Şanlıurfa,Turkey

The aim of this study was to detect the presence of aflatoxin M₁ (AFM1) in samples of raw milk (n = 38), UHT milk (n = 12), white pickled cheese (n = 50), and yogurt (n = 50) collected from the ?anl?urfa city markets and locally produced dairy products by ELISA. The mean contamination rates were 56.74 ± 40.32, 43.1 ± 23.19, 103.2 ± 29.13, and 55.28 ± 12.68 ng/kg, respectively, for raw milk, UHT milk, white pickled cheese, and yogurt. According to the data, 21 (55%) raw milk, 3 (25%) UHT milk, 10 (20%) white pickled cheese, and 10 (20%) yogurt samples were contaminated with AFM1 over the acceptable levels (≥50 ng/kg), ranging from 0.82 to 130.89 ng/kg. None of the white pickled cheese samples contained AFM1 levels above the Turkish legal limit (250 ng/kg). Consequently, the AFM1 contamination levels determined in this study in white pickled cheese were not considered to pose a serious public health hazard. However, the AFM1 levels in raw and UHT milk and yogurt samples indicate an increased human health risk in Turkey related to high aflatoxin levels. Therefore, milk and dairy products have to be monitored by the Turkish public health authorities continuously to detect AFM1 contamination.     

Short communication: Variation in the composition and properties of Swedish raw milk for ultra-high-temperature processing

The composition and properties of raw milk are of great importance for the quality and shelf life of the final dairy product, especially in products with a long shelf life [e.g., ultra-high-temperature (UHT)-treated milk]. The objective of this study was to investigate the compositional variation in raw milk samples before processing at the dairy plant. Moreover, we wanted to investigate the effect of the UHT process on this variation (i.e., if the same variation could be observed in the corresponding UHT milk). The quality traits analyzed included detailed milk composition, counts of total and psychrotrophic bacteria, proteolytic activity, and color, as well as predictive measures of stability (i.e., ethanol stability and heat coagulating time). Samples of raw milk and the corresponding produced UHT milk were collected and analyzed on a monthly basis during 1 yr. Principal component analysis was used to identify months showing similarities and differences with respect to total variation. In contrast to previous studies, we observed only small variations between months and no clear effect of season for the raw milk. For the UHT milk, July and the winter months (December, January, and February) tended to separate from the other months. Quality traits showing significant variation were only to some extent identical in raw milk and UHT-processed milk. A better understanding of the natural variation in raw milk quality will provide opportunities to improve the shelf life of UHT-treated milk products.     

Fluorometric determination of chemically available lysine: adaptation, validation and application to different milk products

A spectrophotometric method based on the reaction between available lysine and ortho-phthaldialdehyde (OPA) was adapted and validated for fluorometric determination of the chemically available lysine contents in milk matrices (UHT and conventional in-bottle sterilized cow milk, milk-based infant formulas and infant formula ingredients). The values of the analytical parameters show its usefulness as a routine method (linearity, r = 0.9992; detection limit, 0.0066 mg/mL assay; accuracy, 99-108%; precision, intra-day 2.1-5.9% and inter-day 3.5 10.2%). No statistically significant differences (p < 0.05) were found between the values obtained with the adapted method and those obtained applying the 1-fluoro-2,4-dinitrobenzene (FDNB) (Carpenter) technique. The OPA method was used to measure the chemically available lysine contents in UHT and sterilized milk marketed in Spain, to study the evolution of chemically available lysine during the shelf-life of UHT milks, and finally the quality of name- and store-brand UHT milks was also compared. No statistically significant differences (p < 0.05) were found between either the available lysine contents of the same type of UHT or sterilized milk or between store- and name-brand UHT milks. Statistically significant differences (p < 0.05) were found between the chemically available lysine contents in UHT and sterilized milk. Losses of chemically available lysine ranging from 2.7 to 29% were obtained during the shelf-life of UHT milk.     

Antioxidant and antigenotoxic effect of dairy products supplemented with red ginseng extract

The purpose of this study was to evaluate the antioxidant and antigenotoxic effect of dairy products milk (M) and yogurt (Y) after the addition of 2% red ginseng extract to milk (RM) and to yogurt (RY). Total phenolic content, total flavonoid content, 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity, oxygen radical absorbance capacity, and total radical trapping antioxidant potential were determined in the samples. Furthermore, antigenotoxic effect of samples was measured, using comet assay in human leukocytes. Total phenolic content and total flavonoid content of RM [38.3 ± 0.8 mg of gallic acid equivalents (GAE)/100 g, 23.6 ± 0.1 mg of quercetin equivalents (QE)/100 g] and RY (41.1 ± 0.9 mg of GAE/100 g, 18.7 ± 0.1 mg of QE/100 g), respectively, were higher than those of M (6.31 ± 0.2 mg of GAE/100 g, 10.4 ± 0.1 mg of QE/100 g) and Y (8.1 ± 0.9 mg of GAE/100 g, 8.4 ± 0.2 mg of QE/100 g), respectively. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity and oxygen radical absorbance capacity values increased significantly after the addition of 2% red ginseng in both. Additionally, the total radical trapping antioxidant potential in RM (787.7 ± 7.0 μg/mL) was lower than in M (2074.0 ± 28.4 μg/mL). The H₂O₂-induced DNA damage in RY (0.1 ± 0.0 mg/mL) was less than the damage in Y (0.4 ± 0.0 mg/mL), but we found no significant difference between M and RM. This study indicates that supplementation with red ginseng can fortify the antioxidant and antigenotoxic effects of dairy products effectively.     

3种鲎试验在乳品内毒素活性定量检测中的应用评价

目的 比较3种内毒素活性定量检测方法在乳品中的应用情况。方法 采用2种传统东方鲎(Tachypleus amebocyte lysate, TAL)试验(动态浊度法/动态显色法)和1种便携式内毒素检测仪(portable test system, PTS)对牛奶、酸奶和奶粉产品内毒素活性进行检测, 评估3种方法在乳品内毒素活性检测中的应用情况。结果 3种方法用于乳品内毒素活性检测的加标回收率为50%~200%, 符合药典要求。对比动态浊度法, 动态显色法检测乳品内毒素活性的加标回收率范围更加接近100%。PTS方法的内毒素活性检测值分别与2种传统TAL试验的检测值有较好的线性关系。结论 3种方法均适用于乳品内毒素活性的检测, 但动态显色法更加适合乳品内毒素的检测。     

液态婴儿奶

液态婴儿奶是供婴儿食用的理想乳制品。目前，国内的婴儿乳制品主要以固态的婴儿奶粉为主，而在国外很多国家（如芬兰、俄罗斯等）液态婴儿奶较流行。液太婴儿奶是超高温（UHT）灭菌处理后经无菌包装而成的产品，产品中不需要添加任何防腐剂，保质期可达6个月以上。与婴儿奶粉相比，液态婴儿奶具有很多优点，如勿需冲调、浓度恒定、喂食方便、安全卫生等。在具体生产过程中，原料和工艺等的选择对最终产品的质量有着很大的影响。     

高效液相色谱法测定婴幼儿配方乳粉中酪蛋白磷酸肽的含量

目的建立高效液相色谱法测定婴幼儿配方乳粉中酪蛋白磷酸肽含量的方法。方法对婴幼儿配方乳粉样品对应生产批次所采用的酪蛋白磷酸肽原料进行测定,得到准确含量后作为参照品,对婴幼儿配方乳粉样品测定前处理中净化、提取条件进行优化,采用二极管阵列检测器(diode array detector, DAD)定量分析。结果采用混合型阳离子交换(mixed cation exchange, MCX)固相萃取柱、ZORBAX Eclipse AAA色谱柱可得到良好的分离效果,酪蛋白磷酸肽在100~500μg/mL范围内线性关系良好,相关系数r^2为0.9994,检出限为0.140 mg/100 g,加标回收为81.06%~87.78%,相对标准偏差为3.64%。结论该方法前处理操作简单,具有良好的重现性和准确性,能够满足婴幼儿配方乳粉中酪蛋白磷酸肽含量的测定。     

湖南省市售国产乳制品中硫氰酸盐检测结果分析

目的掌握湖南省市售国产乳制品中硫氰酸盐含量,有效指导消费者科学消费。方法利用离子色谱法对湖南省388批次市售国产乳制品硫氰酸钠进行检测。结果 388批次乳制品硫氰酸钠检出率为93.3%,折算为生乳后的含量均在国家食药监总局风险监测参考值10 mg/kg范围内。其中,307批次液体乳制品硫氰酸盐检出率为91.5%,按系数折算为生乳后含量在1.0~6.9 mg/kg之间的为86.98%;81批次奶粉检出率为100%,按系数折算为生乳后含量在0.1~5.9 mg/kg之间的为97.35%。结论经过分析原料乳中硫氰酸盐的可能来源,认为湖南省市售国产乳制品中硫氰酸盐主要来源于原料生鲜乳本底,非法添加的可能性较小,有必要立刻全面开展生乳中硫氰酸盐本底值调查,系统地为乳制品硫氰酸盐监测提供有效依据。     

配方注册婴幼儿配方乳粉中蛋白质及三聚氰胺含量测定分析

目的通过测定婴幼儿配方乳粉中蛋白质及三聚氰胺含量,对我国现阶段配方注册的婴幼儿配方乳粉安全状况进行监测。方法婴幼儿配方乳粉中蛋白质的含量依照GB 5009.5-2016《食品安全国家标准食品中蛋白质的测定》中"第一法凯氏定氮法"进行检测;三聚氰胺的含量依照GB/T22388-2008《原料乳与乳制品中三聚氰胺检测方法》中"第一法高效液相色谱法"进行检测。结果样品中蛋白质含量均符合食品安全国家标准规定,且满足测定值不小于标签标示值80%的规定;三聚氰胺的含量均为未检出或低于检出限,符合原卫生部公告的限量值规定。结论根据食品安全国家标准或推荐标准的检测方法和评价指标,所检测的样品中蛋白质项目满足配方注册要求。     

Performance comparison of the BioSys optical assay and the violet red bile agar method for detecting coliforms in food products

Coliform counts in a variety of foods, including dairy products (raw milk, pasteurized milk, yogurt, butter, and ice cream), meats (pork sausage, ground beef, and raw chicken), raw eggs, and chocolate, were performed by the rapid automated BioSys optical assay and the conventional method with violet red bile agar (VRBA). The standard deviation (SD) among five replicate counts for the optical assay was similar to or better than that obtained with VRBA plates for all foods tested. The average SD for all foods tested was 0.21 for the optical assay and 0.30 for the VRBA plates. At very low concentrations of coliforms (1 to 10 CFU/ml for liquid products and 10 to 100 CFU/g for solid samples), the average SDs were 0.26 and 0.47, respectively. The optical assay was less susceptible to interference by noncoliform organisms. In naturally contaminated samples, bacteria such as Serratia liquefaciens, Pantoea spp., Vibrio fluvialis, Aeromonas hydrophilia, and Pseudomonas spp. formed typical colonies in VRBA, resulting in false-positive results or a need to verify colonies in brilliant green lactose broth. The optical assay appeared to be more selective than the VRBA conventional method, detecting fewer noncoliforms. There was close agreement in test results between the two methods, as indicated by correlation coefficients of 0.92 to 0.99 obtained for the regression analysis of the two methods. In most cases both methods distinguished accurately between positive samples containing coliforms and negative controls. All products tested using the automated BioSys Optical Assay for coliforms yielded results more quickly (typically 10 to 12 h) than did those tested with the conventional VRBA method (24 to 72 h with confirmation).     

贮藏期UHT奶中蛋白质组成结构及其糖基化的变化

通过测定UHT奶在贮藏过程中蛋白水解,蛋白在胶束相和乳清相的分布,乳蛋白糖基化位点变化,探究乳蛋白在贮藏过程中性质的变化。结果表明贮藏过程中UHT奶中菌落总数增大,纤溶酶含量增大。UHT奶在内外源酶的作用下乳蛋白发生不同程度的水解。热处理导致从胶束表面脱落的κ-CN和乳清蛋白以热诱导聚合物的形式,在贮藏过程中重新结合到胶束表面,从而使胶束粒径增大,牛乳表观黏度增大。贮藏过程中胶束Zeta电位逐渐降低,流动行为指数n逐渐降低,说明牛乳呈现出更明显的剪切变稀能力,暗示胶束结构更加松散。贮藏过程中UHT奶色泽加深,美拉德反应继续进行,其产物半乳糖基赖氨酸和Nε-羧甲基赖氨酸(CML)含量不断增加。     

Technical note: Development and validation of an HPLC method for the quantification of tocopherols in different types of commercial cow milk

In the present study, a methanol-fluorescence-based HPLC method was validated for its use to quantify α-tocopherol and γ-tocopherol in raw milk, whole UHT milk, partially skimmed UHT milk, whole pasteurized milk, and partially skimmed pasteurized milk. Repeatability and reproducibility, calculated as relative standard deviation of 10 measurements within the same day and 30 measurements across 3 d, respectively, were always below 5% for both tocopherols concentrations and retention times. Recovery was assessed through 3 spiking levels and it ranged from 89 to 107%. The method was able to detect the expected declines in tocopherols in milk exposed to UHT or skimming treatments. Vitamin E, calculated as the sum of α-tocopherol and γ-tocopherol, was similar in whole pasteurized and raw milk, averaging 1.57 and 1.56 mg/L, respectively, followed by whole UHT (1.33 mg/L), partially skimmed pasteurized (0.77 mg/L), and partially skimmed UHT milk (0.61 mg/L).     

The detection of viable vegetative cells of Bacillus sporothermodurans using propidium monoazide with semi-nested PCR

Bacillus sporothermodurans produces highly heat-resistant spores that can survive ultra-high temperature (UHT) treatment in milk. Therefore, we developed a rapid, specific and sensitive semi-nested touchdown PCR assay combined with propidium monoazide (PMA) treatment for the detection of viable B. sporothermodurans vegetative cells. The semi-nested touchdown PCR alone proved to be specific for B. sporothermodurans, and the achieved detection limit was 4 CFU/mL from bacterial culture and artificially contaminated UHT milk. This method combined with PMA treatment was shown to amplify DNA specifically from viable cells and presented a detection limit of 10² CFU/mL in UHT milk. The developed PMA-PCR assay shows applicability for the specific detection of viable cells of B. sporothermodurans from UHT milk. This method is of special significance for applications in the food industry by reducing the time required for the analysis of milk and dairy products for the presence of this microorganism.     

Influence of thermal processing conditions on flavor stability in fluid milk: benzaldehyde

Flavor loss in dairy products has been associated with enzymatic degradation by xanthine oxidase. This study was conducted to investigate the influence of milk thermal processing conditions (or xanthine oxidase inactivation) on benzaldehyde stability. Benzaldehyde was added to whole milk which had been thermally processed at 4 levels: (1) none or raw, (2) high temperature, short time (HTST) pasteurization, (3) HTST pasteurization, additionally heated to 100 degrees C (PAH), and (4) UHT sterilized. Additionally, PAH and UHT milk samples containing benzaldehyde (with and without ferrous sulfate) were spiked with xanthine oxidase. Azide was added as an antimicrobial agent (one additional pasteurized sample without) and the microbial load (total plate count) was determined on d 0, 2, and 6. The concentration of benzaldehyde and benzoic acid in all milk samples were determined at d 0, 1, 2, 4, and 6 (stored at 5 degrees C) by gas chromatography/mass spectrometry in selective ion monitory mode. Over the 6-d storage period, more than 80% of the benzaldehyde content was converted (oxidized) to benzoic acid in raw and pasteurized milk, whereas no change in the benzaldehyde concentration was found in PAH or UHT milk samples. Furthermore, the addition of xanthine oxidase or xanthine oxidase plus ferrous sulfate to PAH or UHT milk samples did not result in benzaldehyde degradation over the storage period.     

Conjugated Linoleic Acid Concentrations in Dairy Products as Affected by Processing and Storage

Total conjugated linoleic acid (CLA) concentrations increased 1.32 fold in salted and 1.27 in unsalted butter but did not alter the ratio of 9-cis 11-trans to total CLA. Nonfat yogurt showed an increase in CLA content with processing (5.25 mg total CLA/g fat) compared to unprocessed raw material (4.40 mg CLA/g fat). No changes in CLA content was observed in processed dairy products such as lowfat yogurt, regular yogurt, lowfat and regular ice cream, sour cream or cheeses such as Mozzarella, Gouda and Cheddar. Storage did not affect CLA concentration in any products suggesting that CLA is a stable component.     

用于PCR检测的乳品中金黄色葡萄球菌DNA提取方法比较研究

比较了乳品中金黄色葡萄球菌DNA的6种提取方法的效果,确定了一种可有效从乳品中提取金黄色葡萄球菌DNA的方法。该方法无需对样品进行增菌,可直接从乳品中提取金黄色葡萄球菌DNA,作为模板进行PCR检测,检测效果良好,对UHT全脂乳、脱脂乳和奶酪的最低检出限分别为10、10CFU/mL和55CFU/g。     

酶联免疫吸附法快速测定不同样品 基质中三聚氰胺

目的 探求不同样品基质中三聚氰胺残留量的快速检测方法.为快速筛查不同种类食品中非法添加三聚氰胺提供技术保障.方法 采用酶联免疫吸附法在奶制品(奶粉、液态奶)、成品饲料(鸡饲料、猪饲料)、饲料原料(鱼粉、肉骨粉、豆粕、麸皮)、肉类(鸡肉、猪肉、内脏)等样品基质中添加一定浓度的三聚氰胺进行测定,并对检测结果进行分析.结果 酶联免疫试剂盒对奶制品和肉类检出限均能达到1.0 mg/kg;对成品饲料基质中的三聚氰胺的检测,检出限可达到2 mg/kg,而对饲料原料中的三聚氰胺的检测,检出限都不能达到2 mg/kg.结论 对奶制品中的三聚氰胺检测完全符合我国的临时限量标准;对成品饲料中的三聚氰胺残留的检测同样符合其限量标准(2.5 mg/kg),而对饲料原料中的三聚氰胺的检测,由于不同基质中检出限不同,不能直接采用酶联免疫法进行快速筛查;酶联免疫法也适用于肉类中的三聚氰胺的检测.