Milk and dairy products are a major source of minerals, particularly calcium, involved in several metabolic functions in humans. Currently, several dairy products are enriched with calcium to prevent osteoporosis. The development of an inexpensive and fast quantitative analysis for minerals is required to offer dairy farmers an opportunity to improve the added value of the produced milk. The aim of this study was to develop 5 equations to measure Ca, K, Mg, Na, and P contents directly in bovine milk using mid-infrared (MIR) spectrometry. A total of 1,543 milk samples were collected between March 2005 and May 2006 from 478 cows during the Walloon milk recording and analyzed by MIR spectrometry. Using a principal component approach, 62 milk samples were selected by their spectral variability and separated in 2 calibration sets. Five outliers were detected and deleted. The mineral contents of the selected samples were measured by inductively coupled plasma atomic emission spectrometry. Using partial least squares combined with a repeatability file, 5 calibration equations were built to estimate the contents of Ca, K, Mg, Na, and P in milk. To assess the accuracy of the developed equations, a full cross-validation and an external validation were performed. The cross-validation coefficients of determination (R 2 cv) were 0.80, 0.70, and 0.79 for Ca, Na, and P, respectively (n = 57), and 0.23 and 0.50 for K and Mg, respectively (n = 31). Only Ca, Na, and P equations showed sufficient R 2 cv for a potential application. These equations were validated using 30 new milk samples. The validation coefficients of determination were 0.97, 0.14, and 0.88 for Ca, Na, and P, respectively, suggesting the potential to use the Ca and P calibration equations. The last 30 samples were added to the initial milk samples and the calibration equations were rebuilt. The R 2 cv for Ca, K, Mg, Na, and P were 0.87, 0.36, 0.65, 0.65, and 0.85, respectively, confirming the potential utilization of the Ca and P equations. Even if new samples should be added in the calibration set, the first results of this study showed the feasibility to quantify the calcium and phosphorus directly in bovine milk using MIR spectrometry.
Cow's milk can be used as a potential source of equol in the human diet. In order to study human intake, however, it is necessary to develop a reliable and sensitive analytical method. This paper reports on the validation of an analytical method using ultra-performance liquid chromatography coupled with a tandem mass spectrometry detector to quantify the equol in commercial milks (raw, whole, semi-skimmed, and skimmed milk). The equol was initially released using enzymatic hydrolysis, and it was then extracted using a double liquid/liquid extraction. The analytical method produced a linear calibration curve with a high correlation coefficient (R 2 ≥0.996) between 5 and 1,000 ng.mL -1 . Good intra-and inter-day precision (≤5.3% and≤5.2%, respectively) and accuracy (≤8.6%) were achieved. The recovery rate differed slightly among the different types of milk, ranging between 60.6± 1.09% and 82.3±5.21%. Good method repeatability was observed (≤15%). There was neither matrix effect nor carry-over effect, and the sample extracts were stable for at least 7 days of storage at -21 °C and 5 °C. The method proved to be specific, sensitive, precise, and accurate and was used for the first time to quantify the equol content in Belgian commercial cow's milk. In all the samples analyzed, equol was present at a concentration ≥10 ng.mL -1 and had a significantly higher content in organic than in conventional milk. The study also found that the mean concentrations of equol were similar for each type of commercial conventional cow's milk.
This paper provides an update and comprehensive review of the analytical methods used for quantifying isoflavones and their metabolites in cow’s milk. Isoflavones are secondary plant metabolites that are similar to 17 β-estradiol in chemical structure. They form one of the most common categories of phytoestrogens. Numerous health benefits have been attributed to isoflavones, but many of these compounds are also considered to be endocrine disruptors, with adverse effects on health. These contradictory trends offer an attractive prospect for future research, and therefore, sensitive and reliable analytical methods are required to clarify various issues about isoflavones. For this review, a structured methodology was used to select 26 relevant articles published between 2005 and 2015 from the Scopus and CAB Abstract databases. The review discusses individual steps of the analytical procedures described in these articles, including sample preparation, instrumental analysis and validation. The most commonly used analytical procedure is sample preparation involving liquid-liquid extraction and an enzymatic hydrolysis step followed by liquid chromatography with mass spectrometry analysis. Currently, however, there is no standardized procedure for the sample preparation and analysis of isoflavones in milk.
A performant method for the simultaneous quantification of daidzein, genistein, formononetin, and biochanin A in forages using an UPLC ® -MS/MS was developed and fully validated. The ultrasound-assisted extraction and enzymatic hydrolysis used in the sample preparation step were optimized using the Box-Behnken experimental design. The optimal extraction conditions used for a representative mix of forage plants were 80 °C, 10 min, and 55 % methanol, and for hydrolysis, they were 20 °C, 18 h, and pH = 6. The chromatographic separation was achieved using an Acquity UPLC ® HSS T3 column, with a water/ methanol linear gradient containing 0.01 % of formic acid at a 0.55 mL min -1 flow rate. The four isoflavones were detected by ESI mass spectrometry in positive ion MRM mode. The method allows high throughput analyses of samples and showed an adequate linear regression model for all isoflavones over a range from 5 to 125 ng mL -1 . There were good intra-and interday precisions (≤8.2 and ≤7.6 %) and accuracy (≤11.4 and ≤7.1 %). The recovery rates were in an acceptable range of 70-120 %, except for biochanin A, where the rate was about 50 %. Good method repeatability was also observed, and there was no matrix effect or carryover problem. The sample extracts were stable for at least 6 days of storage at -21 and 6 °C. The method proved to be sensitive, precise, and accurate for discriminating a wide variety of forages likely to be grazed by ruminants according to their isoflavone contents and to observe the impact of storage process on isoflavone content in forages.formononetin, and biochanin A in forages silages originating from three different meadows. This development is part of a larger study on the metabolization of the biomolecules of interest and the potential accumulation of metabolites such equol in milk [15,16]. Materials and Methods Chemicals and ReagentsA freeze dried forage sample containing six plants (Trifolium pratense L., Trifolium repens L., Medicago sativa L., Lotus corniculatus L., Medicago lupulina L, and Lolium perenne L.; 1:1:1:1:1:1, w/w/w/w/w/w) was mixed with flaxseed meal [4:1, w/w; 93.7 ± 0.1 % dry matter). Formononetin 'FO' (CAS number: 485-72-3), biochanin A 'BA' (491-80-5), genistein 'GE' (446-72-0), and flavone 'IS' (525-82-6) were bought from Sigma-Aldrich (Diegem, Belgium). Daidzein 'DA' (486-66-8) was acquired from Cayman Europe (Tallinn, Estonia). Daidzeind 4 (1219803-57-2) was purchased from C/D/N ISOTOPES (Pointe-Claire, Canada). Individual molecule stock solutions (100 µg mL -1 ) were prepared in methanol and stored at -20 °C in the dark. β-Glucosidase (from almonds, ≥6 U rng -1 , 9001-22-3), β-glucuronidase (type H-2 from Helix pomatia, ≥ 85000 units mL -1 , 9001-45-0), and cellulase (from Aspergillus niger, ≥0.3 units mg -1 , 9012-54-8) were bought from Sigma-Adlrich (Diegem, Belgium). Sodium acetate (127-09-3) was obtained from Merck KGaA (Darmstadt, Germany). Throughout the study, the powder enzymes and Helix pomatia juice were dissolved in different proportions of 0.2-M sodium ace...
-Incidence of complemented feed on false positive results with Delvotest SP® for detection of antibiotics in milk. The study described in this paper was performed to show the influence of feed complemented by minerais, oligo-elements and vitamins (CM V) on the results of Delvotest SP®. Three trials were developed. In the first trial, a group of 13 dairy cows was divided into two subgroups of six and seven cows, receiving the first one a feed basis without CMV, the second one, the same feed basis added of 100 g CMV per day. During the second trial, the dose of CMV was doubled for the last seven cows. During the third trial, ail the 8 cows were grazing. The four cows of the test group received 200 g of CMV per day. The two homogeneous groups of cows (somatic cells, number of lactations) were followed for detection of inhibitors in milk using Delvotest SP®. The test gave positive results with milk from complemented cows during the first and the second trials. Ali results were negative during the third trial. Feed appears to be a factor that may give false positive results in the determination of antibiotics in milk with Delvotest SP®. © Inra/Elsevier, Paris.
The objective of this experiment was to compare the nutritional properties of potato protein concentrate, a by-product of the starch industry produced entirely in Europe, with that of soybean meal (SBM), for growing cattle. The experiment was conducted on double-muscled Belgian Blue bulls, fitted with rumen, duodenal and ileal cannulas, according to a 4 × 4 Latin square design. They were fed three different iso-N and iso-net energy diets formulated according to the Dutch feed evaluation system, differing in the nature of the main protein source, which was either SBM (‘SBM’ treatment), potato protein concentrate (PPC, ‘PPC’ treatment) or an iso-N mixture of these two protein sources (‘mixed’ treatment). A fourth treatment consisted of ‘PPC’ supplemented by 9.5% digestible proteins supplied by duodenal perfusion of sodium caseinate (CAS, ‘PPC + CAS’ treatment). No significant difference was observed in the ruminal fluid pH, whereas both ‘PPC’ and ‘PPC + CAS’ had the effect of reducing the ruminal ammonia nitrogen (N-NH3) concentration. No significant difference was observed in the apparent intestinal digestibility of the dry matter (DM), organic matter (OM) or N. Outflows of non-NH3-N, microbial proteins and dietary proteins from the rumen were similar for ‘PPC’, ‘SBM’ and ‘mixed’, and increased with CAS infusion by 20%, 17% and 27%, respectively. On the basis ofin vivoobservations, the degradability of SBM and PPC proteins was estimated at 0.60 and 0.43, respectively, corresponding to the values quoted in the literature. The supply of digestible essential amino acids (EAA) was significantly greater with ‘PPC + CAS’ and did not differ among ‘SBM’, ‘mixed’ and ‘PPC’. This illustrates the difficulty of altering the amino acid (AA) pattern of digestible protein by the nature of the protein of dietary origin when an animal is fed a high nutritional value diet. N retention was not affected by replacing SBM with PPC, but increased by 10% with CAS infusion. On the basis of the plasma AA pattern, the supply of digestible Met was probably limiting with ‘SBM’, ‘mixed’ and ‘PPC’. The CAS perfusion supplemented all AA, including Met, leading to increased N retention. This improvement was limited, however, and N utilisation remained unchanged between treatments. In conclusion, despite a more favourable EAA pattern, PPC offered no advantage compared with SBM for growing bulls when diets were formulated according to the Dutch feed evaluation system.
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