Prediction of the ripening times of ewe's milk cheese by multivariate regression analysis of capillary electrophoresis casein fractions

The effect of the ripening time on the proteolytic process in cheeses made from ewe's milk during a 139-day ripening period was monitored by the use of capillary electrophoresis of pH 4.6 insoluble fraction. Totals of 18 and 21 peaks were recognized and matched in the electropherograms obtained with a fused-silica capillary and a neutral capillary (hydrophilically coated), respectively. These peaks correspond to intact ovine caseins and their hydrolysis products (alpha(s1)-casein I, alpha(s1)-casein II, alpha(s1)-casein III, alpha(s2)-casein, beta(1)-casein, beta(2)-casein, p-kappa-casein, alpha(s1)-I-casein, gamma(1)-casein, gamma(2)-casein, and gamma(3)-casein). The alpha(s)-caseins (alpha(s1)- and alpha(s2)-casein) displayed similar degradation pattern to one another, but different from those of beta-caseins (beta(1)- and beta(2)-casein). beta-Caseins were very much undergoing lesser degradation during the ripening time than alpha(s)-casein. Finally, partial least-squares regression and principal components regression were used to predict the ripening time in cheeses. The models obtained yielded good results since the root-mean-square error in prediction by cross validation was <8.6 days in all cases.     

Quantification of beta casein in milk and cheese using an optical immunosensor

beta-Casein was quantified in milk and cheese, using an optical immunosensor, based on surface plasmon resonance (SPR) measurement. The assay consists of a two-step sandwich strategy, with two anti-beta-casein antibodies directed against each extremity of the casein. This strategy permits only native beta-casein to be quantified and not its degradation products. The calibration curve was obtained with a reference milk powder of known beta-casein concentration. The analysis time per sample was less than 10 minutes. The antibody-coated surface could be used for more than 250 determinations. The detection limit was established at 85 ng x mL(-)(1) and the intra- and inter-assay variation coefficients were 2.6 and 6.2% respectively. The method was applied to raw milk to quantify intact beta-casein, with no pretreatment of the sample. A second application was realized with cheese, to follow the proteolysis of beta-casein during ripening.     

毛细管电泳在兽医药品分析中的应用

综述了近年来毛细管电泳技术在兽医药品分析中的应用和进展,主要包括抗生素类残留、磺胺类残留、激素及β-兴奋剂类残留等的检测,并对其今后的发展作了展望。     

Micellar electrokinetic capillary electrophoresis for rapid analysis of patulin in apple cider

A micellar electrokinetic capillary chromatography (MECC) mode was applied to a capillary electrophoresis (CE) method, which was developed for detection and quantitation of patulin in apple ciders. This method used a small sample amount (2 mL) and consumed minimal organic solvent compared to the most commonly used HPLC methods. The sample preparation procedure of the CE method was also simpler than other chromatographic techniques developed for patulin analysis. Patulin was detected with a photodiode array detector at 273 nm. The standard curve was linear (r(2) = 0.9984) from 75 microgram/L to 121 microgram/mL with patulin working solutions corresponding to 3.8 microgram/L to 6.1 microgram/mL patulin in the sample. The linearity was better in a narrower range of concentrations (r(2) = 0.9999) from 75 microgram/L to 24.1 microgram/mL. The limit of detection of the method was 3.8 microgram/L. Patulin recoveries at 4 levels in spiked samples (10-121 microgram/L) ranged from 95.2 to 105.4%. The recoveries were 96. 9% and 99.2% for 2 levels (22.3 and 223 microgram/L, respectively) of patulin in infected apple samples. This method represents a unique alternative method for rapid and sensitive analysis of patulin in apple ciders.     

Differential characteristics of fatty acids in cheese from milk of various animal species by capillary gas chromatography

The kind of milk used in the manufacture of cheese has been identified by analysis of the fatty acids. The milk fat is extracted from the cheese and saponified. The methyl esters of the fatty acids are prepared and determined by capillary column gas chromatography. Seven major fatty acids are separated and quantitated, namely, C8:0, C10:0, C12:0, C14:0, C16:0, C18:0, and C18:1. Many of the 21 simple ratios that can be formed from these 7 quantities are characteristic of the type of milk from which the fatty acids were obtained. The method allows the identification of cheese prepared with the milk of cows, buffalo, sheep, or goats. Substitution or adulteration of milk can also be detected.     

Near-infrared analysis of fat, protein, and casein in cow's milk.

Fat, crude protein, true protein, and casein were determined in cow milks by near-infrared transmission spectroscopy (NIR). Partial and overall PLS calibrations were performed on two sets of samples: partial calibration included 76 unhomogenized samples, whereas overall calibration used 96 homogenized and unhomogenized samples. Standard errors of calibration were 0.12% for fat, 0.06% for crude protein, 0.04% for true protein, and 0.05% for casein in the overall calibration. Validation of the overall calibration with an independent set of samples gave standard errors of prediction of 0. 07% for fat, 0.06% for crude protein and casein, and 0.05% for true protein. Except for fat, all of the statistical parameters were better with overall than with partial calibrations, which indicates that homogenization has an effect on NIR fat determination. Despite the relatively small number of samples included in the calibration model, NIR transmission was found to be a reliable method for the determination of fat and nitrogenous constituents in milk.     

Determination of phenolic compounds and ascorbic acid in different fractions of tomato by capillary electrophoresis with electrochemical detection

Tomato (Lycopersicon esculentum Mill.), one of the most important crops worldwide, contains different classes of substances with antioxidant properties such as carotenoids, vitamin C, and phenolics. A method based on capillary electrophoresis with electrochemical detection has been developed to analyze ascorbic acid and phenolics in the peel, pulp, and seeds of tomatoes. Operating in a wall-jet configuration, a 300 microm diameter carbon disk electrode was used as the working electrode, which exhibits a good response at +0.90 V (vs saturated calomel electrode) for the analytes. Under optimum conditions, the analytes were baseline separated within 20 min in a 50 mmol/L borate buffer (pH 8.7). Notably, excellent linearity was obtained over 3 orders of magnitude with detection limits (S/N=3) ranging from 1x10(-8) to 2x10(-7) g/mL for all analytes. This proposed method has been successfully applied to monitor the content of ascorbic acid and phenolics in real samples, and the assay results were satisfactory.     

Authentication of coffee by means of PCR-RFLP analysis and lab-on-a-chip capillary electrophoresis

Coffee is one of the most important world food commodities, commercial trade consisting almost entirely of Arabica and Robusta varieties. The former is considered to be of superior quality and thus attracts a premium price. Methods to differentiate these coffee species could prove to be beneficial for the detection of either deliberate or accidental adulteration. This study describes a molecular genetics approach to differentiate Arabica and Robusta coffee beans. This employs a Polymerase Chain Reaction-Restriction Fragment Length Polymorphism to monitor a single nucleotide polymorphism within the chloroplastic genome. Samples were analyzed with a lab-on-a-chip capillary electrophoresis system. Coffee powder mixtures were analyzed with this technique, displaying a 5% limit of detection. The plastid copy number was found to be relatively constant across a wide range of bean samples, suggesting that this methodology can also be employed for the quantification of any adulteration of Arabica with Robusta beans.     

Quantification of the interactions between beta-lactoglobulin and pectin through capillary electrophoresis analysis

Biopolymer interactions have many potential applications in pharmaceutical, cosmetic, nutraceutical, and functional food industries. Attractive interactions between proteins and polysaccharides can lead to the formation of complexes. Binding parameters of beta-lactoglobulin (beta-lg)/pectin complexes were determined using frontal analysis continuous capillary electrophoresis and the overlapping binding site model. At pH 4, approximately 23 beta-lg molecules were cooperatively complexed on low-methoxyl pectin, where each beta-lg molecule covered an average of 12 galacturonic acid residues. The calculated binding constant was 1431 M(-1). The interactions between pectin and four selected peptides located on the outer surface of the beta-lg were investigated in order to identify which part of the protein was likely to interact with the pectin. The peptide beta-lg 132-148, which corresponds to the alpha-helix zone, and the peptides beta-lg 76-83, 41-60, and 1-14 would be involved in the interaction with the pectin.     

Free-zone capillary electrophoresis analysis of hordein patterns at different stages of barley malting

We have carried out a comparison of hordein patterns at different stages of the malting process using free-zone capillary electrophoresis (FZCE). FZCE has proved to be a suitable technique for the separation and characterization of hordeins in barley seeds. Assays of protein extraction and electrophoretic procedures led us to conclude that hordeins were best extracted with 40% ethanol and analyzed using 50 mM phosphate-glycine, pH 2.5, containing 20% ACN and 0.05% HPMC, at 12.5 kV and 45 degrees C, with 10 s hydrodynamic injection at 0.5 psi and 50 microm i.d. x 31 cm uncoated fused-silica capillary. Our results afford useful information about changes in the composition of these proteins in barley during malting.     

牦牛曲拉干酪素脱色工艺优化

用曲拉生产干酪素的过程中,加碱、加热及原料所含的杂质都会促进干酪素发生美拉德反应,产生黄色或褐色物质,影响干酪素的品质。该研究选用保险粉(低亚硫酸钠)、亚硫酸钠、亚硫酸氢钠3种漂白剂,采用L₂₅(5⁶)正交试验设计,对试验结果进行分析,确定出漂白剂的最佳用量为：每8 g曲拉添加保险粉Ⅰ0.15 g、亚硫酸钠0.4g、亚硫酸氢钠0.375 g、保险粉Ⅱ0.2 g。对曲拉原料、曲拉干酪素、鲜奶干酪素进行红外光谱分析比较,表明曲拉干酪素在风味、有效成分上与鲜奶干酪素相近。     

Extraction parameters and capillary electrophoresis analysis of limonin glucoside and phlorin in citrus byproducts.

Limonin glucoside (LG) and phlorin were extracted from citrus fruit tissues and assayed by capillary electrophoresis (CE). LG was determined in dried [1.20 +/- 0.10 mg of dry weight (dw)] and wet peel residues (1.16 +/- 0.04 mg of dw), orange juice finisher pulp (0.58 +/- 0.03 mg of dw), dried grapefruit seeds (2.70 +/- 0.15 mg of dw), and 50 degrees Brix molasses (2225 +/- 68 mg/L). Phlorin was purified from orange peel residue and grapefruit albedo, and concentrations were determined in some citrus products. Phlorin and LG were extracted from residues with water/pectinase or with water solutions of methanol and ethanol. Efficient LG extraction from grapefruit seeds (2.40 +/- 0.15 mg/g) was achieved with 50-65% methanol, solvent polarity P' approximately equal to 7-8. Extracts were purified and concentrated by adsorptive resins and HPLC to obtain 95% pure compounds of LG and phlorin. CE analysis did not require extract purification beyond filtration. LG and phlorin migrated as anions in electropherograms containing peaks representing other citrus flavonoids and limonoid glucosides.     

New capillary electrophoresis method for the determination of furosine in dairy products

A new capillary electrophoresis (CE) method was established for the quantitative determination of furosine in dairy products. Sample preparation and suitable electrophoretic conditions allowed accurate and reproducible quantitation of furosine in dairy products. Sample preparation consisted of drying hydrolyzed samples, redissolving them in 0.2 M NaOH, and purifying them by solid-phase extraction. The electrophoretic separation was carried out in an uncoated capillary maintained at 30 degrees C using 0.1 M phosphate buffer containing the additive hexadecyl trimethylammonium bromide (HDTAB, 1.2 mM) (pH 7.0) under 10 kV voltage and reverse polarity. Coefficients of variation of less than 2.25% for migration time and 5.80% for peak areas indicated that the technique was reproducible. The calibration curve followed a linear relationship with a highly significant (p < 0.01) coefficient of multiple determination (R (2) = 0.997). The limit of quantitation was 0.5 ppm, a concentration that corresponds to 4.5 mg/100 g of protein in milk samples. Furosine concentration (mg/100 g of protein) ranges of different dairy products (raw, pasteurized, UHT, and evaporated milks and yogurt) agreed with ranges previously reported. Therefore, the CE method presented is a suitable technique for the routine assessment of furosine in dairy products.     

An improved method for the quantitation of flavonoids in Herba Leonuri by capillary electrophoresis

A capillary electrophoresis system coupled with wall-jet amperometric detection was used to determine five flavonoids (kaempferol, rutin, hyperoside, quercitrin, quercetin) in Herba Leonuri. Several important effect factors, such as the pH and concentration of running buffer, separation voltage, injection time, and detection potential, were investigated to acquire the optimum conditions. With 50 mmoL/L Na(2)B(4)O(7)-100 mmoL/L NaH(2)PO(4) buffer (pH=7.50) as the running buffer, the five flavonoids were baseline separated within 15 min in a 60 cm length capillary at a separation voltage of 15 kV. The relationship between peak currents and analyte concentrations was linear over about 2 orders of magnitude, with detection limits (S/N = 3) ranging from 0.03 to 0.08 microg/mL for all analytes. The use of this method for the quantitation of the above flavonoids present in the real sample of Herba Leonuri was reported.     

A capillary electrophoresis laser-induced fluorescence method for analysis of potato glycoalkaloids based on a solution-phase immunoassay. 2. Performance evaluation

Glycoalkaloids (GAs) occur naturally in potatoes and are toxic to humans and animals. The objective of the present study was to evaluate the performance of a solution-phase immunoassay coupled to capillary electrophoresis with laser-induced fluorescence (CE-LIF) detection for the determination of total glycoalkaloids in potatoes. The immunoassay was based on a competition between potato glycoalkaloids and fluorescently labeled solanidine. Reaction products were separated in the capillary zone electrophoresis mode. A calibration curve of signal vs log[GA] was linear from 50 to 400 nM. The CV for duplicate and day-to-day analyses averaged 5.7% and 12%, respectively. Spike recoveries ranged from 85 to 97% for spike levels ranging from 43 to 170 microg/g fresh potato. Potato samples with GA concentrations ranging from <40 to >200 microg/g were successfully analyzed, indicating that immuno-CE-LIF is a rapid alternative to traditional ELISA and HPLC methods.     

High-performance capillary electrophoresis with indirect UV detection for determination of alpha-galactosides in Leguminosae and Brassicaceae

A rapid, easy, and reproducible capillary electrophoresis method for determination of raffinose family oligosaccharides (alpha-galactosides) was developed. Sucrose, raffinose, stachyose, verbascose, and ajugose were determined with indirect UV detection at moderate alkaline pH 9.2, using pyridine-2,6-dicarboxylic acid as background electrolyte in a sodium tetraborate buffer with added cetyltrimethylammonium bromide. The separation efficiency measured by the number of theoretical plates (N) ranged from 1.4 x 10(5) to 2.3 x 10(5). The precision of the method, measured by the relative standard deviation (RSD), was less than 0.53% for the migration times and better than 3.4% for normalized areas (NA), considering all sugars except verbascose (RSD(NA) = 11.8%). Detection limits were about 110 microg/mL, corresponding to 150-320 microM. Relative response factors (RRF) were calculated on the basis of linearity studies and used for quantification of alpha-galactosides in a lupine sample (Lupinus angustifolius).     

A rapid sample screening method for authenticity control of whiskey using capillary electrophoresis with online preconcentration

The present study aimed to develop a methodology using capillary electrophoresis for the determination of sinapaldehyde, syringaldehyde, coniferaldehyde, and vanillin in whiskey samples. The main objective was to obtain a screening method to differentiate authentic samples from seized samples suspected of being false using the phenolic aldehydes as chemical markers. The optimized background electrolyte was composed of 20 mmol L(-1) sodium tetraborate with 10% MeOH at pH 9.3. The study examined two kinds of sample stacking, using a long-end injection mode: normal sample stacking (NSM) and sample stacking with matrix removal (SWMR). In SWMR, the optimized injection time of the samples was 42 s (SWMR42); at this time, no matrix effects were observed. Values of r were >0.99 for the both methods. The LOD and LOQ were better than 100 and 330 mg mL(-1) for NSM and better than 22 and 73 mg L(-1) for SWMR. The CE-UV reliability in the aldehyde analysis in the real sample was compared statistically with LC-MS/MS methodology, and no significant differences were found, with a 95% confidence interval between the methodologies.     

Evaluation of capillary gas chromatography for multicomponent drug analysis

Capillary gas chromatography is evaluated for multicomponent drug analysis. A 0.32 mm id X 30 m fused silica column and a 0.75 mm id X 30 m borosilicate glass column (both with OV-1 bonded phase) are investigated. Retention times (relative to caffeine) are presented for 39 drug components representing a variety of chemical classes and pharmacological activities. Reproducibility data are presented for both isothermal and programmed temperature runs on selected drug mixtures. Recovery data are provided for 2 multicomponent drug mixtures prepared to approximate over-the-counter antihistaminic and antitussive preparations.     

Straightforward process for removal of milk fat globule membranes and production of fat-free whey protein concentrate from cheese whey

A straightforward method for the separation of milk fat globule membrane (MFGM) and production of fat-free whey protein concentrate/isolate from cheese whey has been developed. Lowering of the conductivity of the whey from its initial value of about 5600 μS cm(-1) to about 2000-500 μS cm(-1) via diafiltration with water caused selective precipitation of MFGM when incubated for 30 min at pH 4.2 and 35 °C. The whey proteins remained soluble in the supernatant under these conditions. Experimental evidence suggested that precipitation of MFGM at pH 4.2 was not due to a nonspecific effect of lowering of the conductivity of the whey but due to the specific effect of removal of Ca2+ from the whey. The lipid content of whey protein isolate obtained by this process was <0.2%, and the protein loss was <14%. The method provides an industrially feasible process for the production of fat-free whey protein concentrate/isolate. The MFGM, which is reported to contain bioactive/nutraceutical lipids and proteins, is a valuable byproduct of the process.