Detection of Ovine or Bovine Milk Components in Commercial Camel Milk Powder Using a PCR-Based Method

Food ingredient adulteration, especially the adulteration of milk and dairy products, is one of the important issues of food safety. The large price difference between camel milk powder, ovine, and bovine milk powder may be an incentive for the incorporation of ovine and bovine derived foods in camel milk products. This study evaluated the use of ordinary PCR and real-time PCR for the detection of camel milk powder adulteration based on the presence of ovine and bovine milk components. DNA was extracted from camel, ovine, and bovine milk powder using a deep-processed product column DNA extraction kit. The quality of the extracted DNA was detected by amplifying the target sequence from the mitochondrial Cytb gene, and the extracted DNA was used for the identification of milk powder based on PCR analysis. In addition, PCR-based methods (both ordinary PCR and real-time PCR) were used to detect laboratory adulteration models of milk powder using primers targeting mitochondrial genes. The results show that the ordinary PCR method had better sensitivity and could qualitatively detect ovine and bovine milk components in the range of 1% to 100% in camel milk powder. The commercial camel milk powder was used to verify the practicability of this method. The real-time PCR normalization system has a good exponential correlation (R² = 0.9822 and 0.9923) between ovine or bovine content and Ct ratio (specific/internal reference gene) and allows for the quantitative determination of ovine or bovine milk contents in adulterated camel milk powder samples. Accuracy was effectively validated using simulated adulterated samples, with recoveries ranging from 80% to 110% with a coefficient of variation of less than 7%, exhibiting sufficient parameters of trueness. The ordinary PCR qualitative detection and real-time PCR quantitative detection method established in this study proved to be a specific, sensitive, and effective technology, which is expected to be used for market detection.     

Detection of royal jelly adulteration using carbon and nitrogen stable isotope ratio analysis

Stable isotope ratios ((13)C/(12)C and (15)N/(14)N) were measured in royal jelly (RJ) samples by isotope ratio mass spectrometry (IRMS) to evaluate authenticity and adulteration. Carbon and nitrogen isotope contents (given as delta values relative to a standard, delta(13)C, delta(15)N) of RJ samples from various European origins and samples from commercial sources were analyzed. Uniform delta(13)C values from -26.7 to -24.9 per thousand were observed for authentic RJ from European origins. Values of delta(15)N ranged from -1.1 to 5.8 per thousand depending on the plant sources of nectars and pollen. High delta(13)C values of several commercial RJ samples from -20.8 to -13.3 per thousand indicated adulteration with high fructose corn syrup (HFCS) as a sugar source. Use of biotechnologically produced yeast powder as protein source for the adulterated samples was assumed as delta(15)N values were lower, as described for C(4) or CAM plant sources. RJ samples from authentic and from adulterated production were distinguished. The rapid and reliable method is suitable for urgent actual requirements in food monitoring.     

Rapid impedance method for predicting the potential shelf life of packaged pasteurized milk

The potential shelf life of packaged pasteurized milk was studied using a rapid impedance method. The results showed a high correlation between the shelf life of Fengxing and Xiang Man-lou pasteurized milks and between the detection time and the logarithm of colony-forming units per milliliter. The impedance detection time was measured after preliminary incubation at 30 degrees C for 6 h for 100 and 200 microL milk samples, or at 37 degrees C for 6 h for 100-400 microL milk samples for Fengxing pasteurized milk, and after 6 h preliminary incubation at 30 degrees C for 300 and 400 microL milk samples or 6 h at 37 degrees C for 100-400 microL milk samples for Xiang Man-lou pasteurized milk. Regressive equations were then constructed to predict the potential shelf life. Compared with the traditional method, the impedance method plus pre-incubation of milk at elevated temperatures (30 and 37 degrees C) was rapid, accurate, and convenient. The entire estimation process was completed within 11-14 and 14-20 h for Fengxing and Xiang Man-lou pasteurized milks, respectively.     

Determination of vegetal proteins in milk powder by sodium dodecyl sulfate-capillary gel electrophoresis: interlaboratory study

An interlaboratory study, with the participation of 8 laboratories, was conducted to evaluate a sodium dodecyl sulfate-capillary gel electrophoresis method for determination of adulteration of milk powder with soy and pea proteins. Calibration standards (0-8%, w/w, soy and pea protein in total protein) and adulterated skim milk powders (0-5%, w/w, soy and pea proteins in total protein) were produced. Vegetal proteins were determined after removal of milk proteins by pretreatment of the samples with tetraborate-EDTA buffer, pH 8.3. Repeatability standard deviations ranged from 9 to 15% and reproducibility standard deviations ranged from 25 to 30% in the samples containing 5% vegetal protein in total protein.     

A novel strategy to verification of adulteration in alcoholic beverages based on Schlieren effect measurements and chemometric techniques

A novel strategy to evaluation of adulteration in alcoholic beverages based on the measurement of the Schlieren effect using an automated FIA system with photometric detection is proposed. The assay is based on the Schlieren effect produced when beverage samples are injected in a single-line FIA system that uses water as carrier stream and a light-emitting diode-phototransistor photometer controlled by microcomputer as detector. The flow system presents limited mixing conditions which make possible to create gradients of refractive index (Schlieren effect) in the injected sample zone. These gradients are reproducible, characteristic of each alcoholic beverage and undergo specific modifications when adulterations with water or ethanol are imposed. Schlieren effect data of brandies, cachaças, rums, whiskies and vodkas were treated by SIMCA to elaborate class models applied in the evaluation of alcoholic beverages adulteration. Samples of the original matrix of each sort of beverages were adulterated in laboratory by adding water, methanol and ethanol in levels of 5% and 10% (v/v). These samples were used as test set to validate SIMCA class models. The verification of authenticity using Schlieren effect measurements presented good results making possible to identify 100% of the beverages samples adulterated in laboratory and 93% of the actual adulterated alcoholic beverages with confidence levels of 95%. As principal advantage, the automated system does not use reagents to carry out the analysis.     

毛细管电泳检测奶粉中添加的大豆分离蛋白

采用毛细管电泳方法对乳蛋白和大豆分离蛋白进行检测。选择聚丙烯酰胺涂层毛细管,在紫外检测波长214 nm、分离电压25 kV条件下测出五种乳蛋白在不同线性范围内的线性相关系数均大于0.998,各乳蛋白的迁移时间和峰面积的相对标准偏差(RSD)分别小于0.34%和4.50%,奶粉样品加标回收率范围为88.07%~102%;通过对比大豆7S和11S组分、大豆分离蛋白及乳蛋白的电泳图谱,定性确定出大豆分离蛋白8个特征峰,各特征峰的迁移时间和峰面积的RSD分别小于0.36%和4.87%,方法重现性较好。建立了奶粉中添加大豆分离蛋白的半定量检测方法。     

Detection and quantification of honey adulteration via direct incorporation of sugar syrups or bee-feeding: preliminary study using high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and chemometrics

Honey adulteration is a complex problem which currently has a significant economic impact and undeniable nutritional and organoleptic consequences. This paper describes the development of an effective anionic chromatographic method (HPAEC-PAD) for honey analysis and adulteration detection. The method relies on the use of chemometric methods to process chromatograms in order to achieve a better discrimination between authentic and adulterated honeys by linear discriminant analysis and to quantify adulteration levels by partial least squares analysis. This approach was investigated using honey samples adulterated from 10 to 40% with various industrial bee-feeding sugar syrups. Good results were obtained in the characterization of authentic and adulterated samples (96.5% of good classification) using linear discriminant analysis followed by a canonical analysis. The application of the partial least squares modeling method provided a corresponding linear regression model allowing the percentage of adulteration of new samples to be estimated directly from sample chromatograms.Additionally, a bee-feeding experiment on a small apiary was conducted in order to evaluate the effect of supplying hives with bee-feeding syrups. This practice is specific to the apicultural area. It has been demonstrated that bee-feeding can modify the sugar composition of the produced honey if it is conducted without safeguards.     

Screening of adulteration in packaging biocomposites by infrared spectroscopy and chemometrics

Fourier transform infrared spectroscopy coupled with chemometrics was employed to detect packaging polylactic acid-based biocomposite samples adulterated with polypropylene (PP) 30–45% and linear low-density polyethylene 2–10%. Principal component analysis, soft independent modeling of class analogy (SIMCA) and partial least square discriminate analysis (PLS-DA) chemometric techniques were utilized to classify samples in different classes. Totally, 362 samples were modeled in three different classes (two adulterated and one non-adulterated). The obtained results revealed that PLS-DA is the most suitable chemometric approach for prediction of probable adulteration in biocomposite samples with reliable specificity and selectivity. It could provide 99% correct class prediction rate between non-adulterated biocomposite samples and adulterated ones, while SIMCA methods provided 73.33% prediction accuracy in classification.     

GC Fingerprints Coupled to Pattern-Recognition Multivariate SIMCA Chemometric Analysis for Brazilian Gasoline Quality Studies

ASTM D6729 gas chromatographic fingerprinting coupled to pattern-recognition multivariate soft independent modeling of class analogy (SIMCA) chemometric analysis provides an original and alternative approach to screening Brazilian commercial gasoline quality. SIMCA, was performed on gas chromatographic fingerprints to classify the quality of representative commercial gasoline samples selected by hierarchical cluster analysis and collected over a 5 month period from gas stations in São Paulo State, Brazil. Following an optimized ASTM D6729 gas chromatographic-SIMCA algorithm, it was possible to correctly classify the majority of commercial gasoline samples. The method could be employed for rapid monitoring to discourage adulteration.     

Rapid,non-destructive determination of butter adulteration by means of photopyroelectric (PPE) calorimetry

The study focuses on the application of the photopyroelectric (PPE) technique combined with gas chromatography to detect adulteration of cow milk-obtained butter. The thermal diffusivity and effusivity have been directly measured using back and front PPE detection, respectively, and the results have been correlated with the composition of adulterated butter samples. The back detection configuration has been used in the case of butter adulterated with palm oil, and a possible correlation of the thermal diffusivity with total amount of monounsaturated fatty acids composition has been proposed. For butter adulterated with soy milk, we used the front PPE configuration in order to measure the samples’ thermal effusivity. A strong dependence of the value of thermal effusivity as a function of soy milk content was found.     

Sequential (step-by-step) detection,identification and quantitation of extra virgin olive oil adulteration by chemometric treatment of chromatographic profiles

An analytical method for the sequential detection, identification and quantitation of extra virgin olive oil adulteration with four edible vegetable oils--sunflower, corn, peanut and coconut oils--is proposed. The only data required for this method are the results obtained from an analysis of the lipid fraction by gas chromatography-mass spectrometry. A total number of 566 samples (pure oils and samples of adulterated olive oil) were used to develop the chemometric models, which were designed to accomplish, step-by-step, the three aims of the method: to detect whether an olive oil sample is adulterated, to identify the type of adulterant used in the fraud, and to determine how much aldulterant is in the sample. Qualitative analysis was carried out via two chemometric approaches--soft independent modelling of class analogy (SIMCA) and K nearest neighbours (KNN)--both approaches exhibited prediction abilities that were always higher than 91% for adulterant detection and 88% for type of adulterant identification. Quantitative analysis was based on partial least squares regression (PLSR), which yielded R2 values of >0.90 for calibration and validation sets and thus made it possible to determine adulteration with excellent precision according to the Shenk criteria.     

Feasibility of near-infrared spectroscopy to detect and to quantify adulterants in cow milk.

Cow milk adulteration involves the dilution of milk with a less-expensive component, such as water or whey. Near-infrared spectroscopy (NIRS) was employed to detect the adulterations of milk, non-destructively. Two adulteration types of cow milk with water and whey were prepared, respectively. NIR spectra of milk adulterations and natural milk samples in the region of 1100 - 2500 nm were collected. The classification of milk adulterations and natural milk were conducted by using discriminant partial least squares (DPLS) and soft independent modelling of class analogy (SIMCA) methods. PLS calibration models for the determination of water and whey contents in milk adulteration were also developed, individually. Comparisons of the classification methods, wavelength regions and data pretreatments were investigated, and are reported in this study. This study showed that NIR spectroscopy can be used to detect water or whey adulterants and their contents in milk samples.     

Lactosylated casein phosphopeptides as specific indicators of heated milks

Casein phosphopeptides (CPP) were identified in small amounts in milks heated at various intensities by using matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry. CPP selectively concentrated on hydroxyapatite (HA) were regenerated using phosphoric acid mixed in the matrix. Unphosphorylated peptides not retained by HA were removed by buffer washing. This procedure enhanced the MALDI signals of CPP that are ordinarily suppressed by the co-occurrence of unphosphorylated peptides. CPP, belonging to the β-casein (CN) family, i.e., (f1-29) 4P, (f1-28) 4P, and (f1-27) 4P, and the α_s2-CN family, i.e., (f1-21) 4P and (f1-24) 4P, were observed in liquid and powder milk. The lactosylated counterparts were specific to intensely heated milks, but absent in raw and thermized/pasteurized milk. Most CPP with C-terminal lysines probably arose from the activity of plasmin; an enzyme most active in casein hydrolysis. A CPP analogue was used as the internal standard. The raw milk signature peptide β-CN (f1-28) 4P constituted ~4.3% of the total β-CN. Small amounts of lactosylated peptides, which varied with heat treatment intensity, were detected in the milk samples. The limit of detection of ultra-high-temperature milk adjunction in raw or pasteurized milk was ~10%.     

低场核磁共振结合化学计量学方法快速检测掺假核桃油

以掺假核桃油样品为低场核磁共振检测对象,利用主成分分析法(PCA)和偏最小二乘回归法(PLSR)分析处理Carr-Purcell-Meiboom-Gill(CPMG)序列的核磁共振弛豫数据,旨在探求一种能快速检测核桃油品质的新方法。对几种常见掺假形式(掺入大豆油、玉米油、葵花油)的核桃油样品和纯核桃油样品进行检测和评价。实验结果表明:纯核桃油和掺入不同种类食用油的掺假核桃油在主成分得分图上可以得到很好的区分,且掺假样品随掺假比例在图中呈规律性分布;采用PLSR法对CPMG数据和实际掺假率进行回归,可实现对核桃油掺假水平的准确定量测定。方法快速、无损、准确,在食用油制品的品质控制及评价方面具有很大的应用潜力。     

Screening Brazilian commercial gasoline quality by hydrogen nuclear magnetic resonance spectroscopic fingerprintings and pattern-recognition multivariate chemometric analysis

The identification of gasoline adulteration by organic solvents is not an easy task, because compounds that constitute the solvents are already in gasoline composition. In this work, the combination of Hydrogen Nuclear Magnetic Resonance (¹H NMR) spectroscopic fingerprintings with pattern-recognition multivariate Soft Independent Modeling of Class Analogy (SIMCA) chemometric analysis provides an original and alternative approach to screening Brazilian commercial gasoline quality in a Monitoring Program for Quality Control of Automotive Fuels. SIMCA was performed on spectroscopic fingerprints to classify the quality of representative commercial gasoline samples selected by Hierarchical Cluster Analysis (HCA) and collected over a 6-month period from different gas stations in the São Paulo state, Brazil. Following optimized the ¹H NMR-SIMCA algorithm, it was possible to correctly classify 92.0% of commercial gasoline samples, which is considered acceptable. The chemometric method is recommended for routine applications in Quality-Control Monitoring Programs, since its measurements are fast and can be easily automated. Also, police laboratories could employ this method for rapid screening analysis to discourage adulteration practices.     

Study on the Performance of Three Different Capillary Gas Chromatographic Analyses in the Evaluation of Milk Fat Purity

The adulteration of milk fat with foreign fat has been and still is a major concern in the dairy industry. Milk fat purity is currently evaluated by triglyceride analysis by using the Official EU method. The detection limit of the various vegetable and animal fats ranges between 4 and 6%. This research was carried out to verify whether it is possible to decrease the detection limits of beef tallow, which is the most widely used adulterating animal fat. For this purpose, determinations of diglycerides and 3,5-cholestadiene, together with the Official EU method, were applied both to several samples of pure milk fat and to mixtures of milk fat with different percentages of beef tallow. The best results were obtained combining the data deriving from the three determinations by multivariate statistical techniques; in particular, the statistical model obtained by the UNEQ technique seems to be able to decrease the detection limit of beef tallow from 5.2 to 2%. The diglyceride and 3,5-cholestadiene evaluation, combined with the Official EU method for triglycerides, can be usefully applied both to detect small additions of beef tallow and to demonstrate the adulteration of milk fat samples showing results close to the detection limit of the official method.     

Determination of cows' milk in goats' milk and cheese by capillary electrophoresis of the whey protein fractions.

The use of capillary zone electrophoresis to determine the adulteration of cows' milk in goats' milk products is described. The detection and quantification of cows' milk was based on the presence of the specific whey proteins: the relative calibration curve is reported. The peaks of interest were well resolved by using sodium borate at pH 9.2 as background electrolyte in methyl-silanized capillaries. The minimum amount detectable of cows' milk was 2% in milk mixtures and 4% in cheeses. Restrictions due to genetic variability and possible heat treatments, on only one of the two types of milk employed, are taken into account. Qualitative analysis of goat-ewe-cow and goat-ewe samples are also reported.     

Capillary electrophoresis of small anions in complex samples with a hollow fiber sampling inlet end.

A hollow fiber sampling inlet end of capillary electrophoresis (CE) for small ions in complex samples is described. A short piece of hollow fiber, which only permits small ions to pass through its wall, was directly connected to one end of a capillary. Small ions in complex samples were injected into the capillary through the hollow fiber by an electrokinetic method. The experimental results of CE for standard anion mixture solutions, river water, and milk samples show that the hollow fiber sampling inlet end is a simple and useful method for the CE of complex samples. Furthermore, it has been shown that the method can be used to study small anion changes during the milk storing process.     

Determination of short-chain free fatty acids in lipolyzed milk fat by capillary electrophoresis

The objective of this study was to monitor the release of short-chain free fatty acids (FFA) from milk fat during hydrolysis with lipase using capillary electrophoresis. Sample and run buffer allowed FFA to be maintained in solution by using cyclodextrin and methanol. Indirect UV detection at 270 nm was used, employing p-anisate as a chromophore. Calibration curves constructed for each individual FFA followed linear relationships with highly significant (p < 0.01) correlation coefficients. Electrophoretic FFA profiles of fresh milk fat and lipolyzed milk fat showed marked qualitative and quantitative differences. Butanoic acid (C4) was found in a concentration of 64 ppm, while hexanoic (C6) and octanoic (C8) acids were found in concentrations of 3.8 ppm in fresh milk fat. After a 60-min hydrolysis with commercial lipase, FFA released from milk fat consisted mainly of high concentrations (ppm) of butanoic (C4) (900), followed by hexanoic (C6) (427), octanoic (C8) (282), decanoic (C10) (92), pentanoic (C5) (47), and dodecanoic (C12) (37.5) acids. Ratios of FFA that were associated with flavor balance were calculated. The application of CE for lipolysis monitoring in milk fat offers a simple and fast method for the determination of FFA. Quantitative data can be obtained in 20 min, including sample preparation. The lengthy and laborious steps required in traditional chromatographic techniques, such as lipid extraction, FFA isolation, and derivatization, were not required in this CE method. The implementation of CE for milk fat lipolysis monitoring may be a useful quality control tool for dairy flavor development and production.     

Carbohydrate specification limits for the authenticity assessment of soluble (instant) coffee: statistical approach

Carbohydrates are very good tracers for the authenticity assessment of soluble (instant) coffee. Total glucose and total xylose were determined in 1002 samples of commercial soluble coffees using high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) according to AOAC Method 995.13 and ISO standard 11292. The samples analyzed included both authentic and adulterated coffees but were all sold as 100% pure soluble coffee. The statistical model, called "mixture of distributions," was applied to all results and allowed to derive the maximum content of the 2 carbohydrates in the population of products corresponding to pure soluble coffee: 2.32% for total glucose and 0.42% for total xylose. The specification limits, above which a soluble coffee should be considered as adulterated, were calculated by adding the corresponding expanded measurement uncertainty to the maximum content. Specification limits of 2.46% for total glucose and 0.45% for total xylose were obtained. The data statistically confirm previously established criteria for the authenticity assessment of commercial soluble coffee.