Total phenolic compounds in milk from different species. Design of an extraction technique for quantification using the Folin-Ciocalteu method.

Milk protects the health of newborns because it contains essential compounds that perform metabolic activities. Despite these benefits, the study of phenolic compounds in milk has been poorly explored. The objective of this study was to develop and validate a technique for extracting total phenolic compounds (TPCs) from a milk matrix and then analyzing them using the Folin-Ciocalteu method. The extraction technique was applied to goat milk and involved the addition of methanol, acetonitrile, and Carrez I and II reagents, after which protein was separated from fat through centrifugation. Subsequently, the technique was applied to goat (69.03±6.23mg GAE/L), cow (49.00±10.77mg GAE/L), sheep (167.6±58.77mg GAE/L) and human milk (82.45±12.3mg GAE/L). The technique showed an acceptable linearity (R(2)=0.9998), limit of detection (6.03mg GAE/L) and quantification (16.2mg GAE/L), repeatability (RSD=4%), reproducibility (RSD=6.8%) and recovery (>85.41%); it is thus effective and can be used in the routine analysis of milk. TPCs obtained from each type of milk indicate a high variability among species and among members of the same species.

Content and evolution of potential furfural compounds in commercial milk-based infant formula powder after opening the packet.

Potential furfural compounds were examined by RP-HPLC-DAD in 20 commercial milk-based powdered infant formula (IF) brands from local markets from Paris, France; DF, Mexico; Copenhagen, Denmark; England, UK; and Barcelona, Spain. We traced the evolution of these compounds after the packets had been opened at 0, 30 and 70 days of storage at room temperature (≈25 °C; minimum 23 °C and maximum 25.5 °C). All formula brands were analysed during the first 3-5 months of their shelf life. The mean values of all IFs for potential 5-hydroxymethyl-2-furaldehyde (HMF)+2-furaldehyde (F) were 1115.2 µg/100 g (just opened), 1157.6 µg/100 g (30 days) and 1344.5 µg/100 g of product (70 days). In general, slight increases of potential furfural contents were observed in most of the studied IFs, which suggests that the Maillard reaction increases after opening the packets. The main furfural compound found was HMF, as expected. The range of potential HMF consumed for an infant about 6 months old feeding only on formula was estimated between 0.63 mg and 3.25 mg per day.

Elevated circulating LDL phenol levels in men who consumed virgin rather than refined olive oil are associated with less oxidation of plasma LDL.

In human LDL, the bioactivity of olive oil phenols is determined by the in vivo disposition of the biological metabolites of these compounds. Here, we examined how the ingestion of 2 similar olive oils affected the content of the metabolic forms of olive oil phenols in LDL in men. The oils differed in phenol concentrations as follows: high (629 mg/L) for virgin olive oil (VOO) and null (0 mg/L) for refined olive oil (ROO). The study population consisted of a subsample from the EUROLIVE study and a randomized controlled, crossover design was used. Intervention periods lasted 3 wk and were preceded by a 2-wk washout period. The levels of LDL hydroxytyrosol monosulfate and homovanillic acid sulfate, but not of tyrosol sulfate, increased after VOO ingestion (P < 0.05), whereas the concentrations of circulating oxidation markers, including oxidized LDL (oxLDL), conjugated dienes, and hydroxy fatty acids, decreased (P < 0.05). The levels of LDL phenols and oxidation markers were not affected by ROO consumption. The relative increase in the 3 LDL phenols was greater when men consumed VOO than when they consumed ROO (P < 0.05), as was the relative decrease in plasma oxLDL (P = 0.001) and hydroxy fatty acids (P < 0.001). Plasma oxLDL concentrations were negatively correlated with the LDL phenol levels (r = -0.296; P = 0.013). Phenols in LDL were not associated with other oxidation markers. In summary, the phenol concentration of olive oil modulates the phenolic metabolite content in LDL after sustained, daily consumption. The inverse relationship of these metabolites with the degree of LDL oxidation supports the in vivo antioxidant role of olive oil phenolics compounds.

Presence of virgin olive oil phenolic metabolites in human low density lipoprotein fraction: determination by high-performance liquid chromatography-electrospray ionization tandem mass spectrometry.

The biological benefits of olive oil in preventing the oxidation of low density lipoprotein (LDL) would seem to be linked to its high monounsaturated fatty acid contents, but also to its respective phenolic compounds contents. One prerequisite to assess the in vivo physiological significance of phenolic compounds is to determine their presence in human LDL following the ingestion of virgin olive oil. In this work, olive oil phenolic metabolites were identified using high-performance liquid chromatography in tandem with electrospray mass spectrometry (HPLC-ESI-MS/MS) detection, after solid phase extraction (SPE). Quantitative methods were developed in carrying out linearity, precision, sensitivity and recovery tests. The results from two methods of LDL separation were compared and shorter LDL isolation procedure showed a better recovery for antioxidants compounds in LDL. The metabolites identified in LDL were: hydroxytyrosol monoglucuronide, hydroxytyrosol monosulfate, tyrosol glucuronide, tyrosol sulfate and homovanillic acid sulfate. The fact that olive oil phenolic metabolites are able to bind LDL strengthens claims that these compounds act as in vivo antioxidants.

Simultaneous analysis of Vitamins A and E in infant milk-based formulae by normal-phase high-performance liquid chromatography-diode array detection using a short narrow-bore column.

A rapid, simple and reproducible normal-phase (NP) high-performance liquid chromatography (HPLC)-diode array detection (DAD) method for simultaneous qualitative and quantitative determination of Vitamin A (retinol acetate and retinol palmitate) and Vitamin E (alpha-tocopherol acetate, alpha-, gamma- and delta-tocopherols) in milk-based infant formulae was developed and validated. The preparation sample was based on protein precipitation and vitamin extraction with ethanol, followed by re-extraction with hexane, while the chromatographic method was based on the use of a short narrow-bore column (50 mm x 2.1 mm; 3 microm particle size), which afforded less solvent consumption and higher mass sensitivity. The method showed acceptable values for precision, recovery and sensitivity, and proved very simple for routine analysis work.

Analysis of potential and free furfural compounds in milk-based formulae by high-performance liquid chromatography. Evolution during storage.

A simple and reproducible HPLC-diode array detection method for the qualitative and quantitative analysis of potential and free furfural compounds (5-hydroxymethyl-2-furaldehyde, HMF; 2-furaldehyde, F; 2-furyl methyl ketone, FMC; and 5-methyl-2-furaldehyde, MF) in milk-based formulae was developed and validated. The method showed good linearity with determination coefficients over 0.999. The limits of detection and quantification were acceptable for all furfurals. The relative standard deviations (RSDs) for repeatability and reproducibility were <4.28. Recoveries in all furfurals were between 94.5 and 98.7%. In addition, we report the evolution over shelf life of furfural compound levels in an experimental powder formula for pregnant women stored at 25 and 37 degrees C from production until 15 months.

Analysis of mono- and disaccharides in milk-based formulae by high-performance liquid chromatography with refractive index detection.

A simple and reproducible method for the qualitative and quantitative analysis of free mono- and disaccharides (fructose, glucose, galactose, sucrose, lactulose and lactose) in milk-based formulae by high-performance liquid chromatography (HPLC) with refractive index (RI) detection was developed and validated. The method showed good linearity with determination coefficients exceeding 0.99. The limits of detection (DL) in these sugars were 0.17, 0.13, 0.06, 0.16, 0.05 and 0.25 mg/ml, respectively; and the limits of quantification (QL), 0.27, 0.24, 0.20, 0.26, 0.22 and 0.38 mg/ml. The relative standard deviations (R.S.D.s) for repeatability in fructose, sucrose, lactulose and lactose were 0.78, 0.99, 2.91 and 0.46 and the R.S.D.s for reproducibility were 4.8, 6.15, 7.04 and 2.49, respectively. Recoveries in all sugars were between 93 and 113%.