Effect of dietary α-tocopherol on the bioavailability of lutein in laying hen.

Lutein and its isomer zeaxanthin have gained considerable interest as possible nutritional ingredient in the prevention of age-related macular degeneration (AMD) in humans. Egg yolk is a rich source of these carotenoids. As an oxidative sensitive component, antioxidants such as α-tocopherol (T) might contribute to an improved accumulation in egg yolk. To test this, chickens were fed lutein esters (LE) with and without α-tocopherol as an antioxidant. After depletion on a wheat-soya bean-based lutein-poor diet for 21 days, laying hens (n = 42) were equally divided into three groups and fed the following diets for 21 days: control (basal diet), a LE group (40 mg LE/kg feed) and LE + T group (40 mg LE plus 100 mg T/kg feed). Eggs and blood were collected periodically. Carotenoids and α-tocopherol in yolk and blood plasma were determined by HPLC. Egg yolk was also analysed for total carotenoids using a one-step spectrophotometric method (iCheck((™)) ). Lutein, zeaxanthin, α-tocopherol and total carotenoids in egg yolk were highest after 14 days of feeding and decreased slightly afterwards. At the end of the trial, eggs of LE + T group contained higher amount of lutein (13.72), zeaxanthin (0.65), α-tocopherol (297.40) and total carotenoids (21.6) compared to the LE group (10.96, 0.55, 205.20 and 18.0 mg/kg, respectively, p < 0.05). Blood plasma values of LE + T group contain higher lutein (1.3), zeaxanthin (0.06) and tocopherol (20.1) compared to LE group (1.02, 0.04 and 14.90 mg/l, respectively, p < 0.05). In conclusion, dietary α-tocopherol enhances bioavailability of lutein reflecting higher content in egg yolk and blood plasma. Improved bioavailability might be due to increased absorption of lutein in the presence of tocopherol and/or a greater stability of lutein/zeaxanthin due to the presence of α-tocopherol as an antioxidant.

Influence of different oral rehydration solutions on abomasal conditions and the acid-base status of suckling calves.

The aim of the study was to investigate the influence of oral rehydration solutions (ORS) on milk clotting, abomasal pH, electrolyte concentrations, and osmolality, as well as on the acid-base status in blood of suckling calves, as treatment with ORS is the most common therapy of diarrhea in calves to correct dehydration and metabolic acidosis. Oral rehydration solutions are suspected to inhibit abomasal clotting of milk; however, it is recommended to continue feeding cow's milk or milk replacer (MR) to diarrheic calves to prevent body weight losses. Three calves with abomasal cannulas were fed MR, MR-ORS mixtures, or water-ORS mixtures, respectively. Samples of abomasal fluid were taken before and after feeding at various time points, and pH, electrolyte concentrations, and osmolality were measured. The interference of ORS with milk clotting was examined in vivo and in vitro. To evaluate the effects of ORS on systemic acid-base status, the Stewart variables strong ion difference ([SID]), acid total ([A(tot)]), and partial pressure of CO2 (pCO2) were quantified in venous blood samples drawn before and after feeding. Calves reached higher abomasal pH values when fed with MR-ORS mixtures than when fed MR. Preprandial pH values were re-established after 4 to 6 h. Oral rehydration solutions prepared in water increased the abomasal fluid pH only for 1 to 2 h. Oral rehydration solutions with high [SID(3)] ([Na(+)] + [K(+)] - [Cl(-)]) values produced significantly higher abomasal pH values and area under the curve data of the pH time course. Caseinomacropeptide, an indicator of successful enzymatic milk clotting, could be identified in every sample of abomasal fluid after feeding MR-ORS mixtures. The MR-ORS mixtures with [SID(3)] values > or =92 mmol/L increased serum [SID(3)] but did not change venous blood pH. Oral rehydration solutions do not interfere with milk clotting in the abomasum and can, therefore, be administered with milk. In this study, MR-ORS mixtures with high [SID(3)] values caused an increase of serum [SID(3)] in healthy suckling calves and may be an effective treatment for metabolic acidosis in calves suffering from diarrhea.

Interactions of glycinin with plant phenols--influence on chemical properties and proteolytic degradation of the proteins.

Soya glycinin was derivatized with different phenolic substances (caffeic-, chlorogenic-, gallic acid and quercetin). The protein derivatives formed have been characterized in terms of their properties where they showed changes in the content of free epsilon-amino groups, tryptophan and thiol groups. The derivatives have also been characterized in terms of their solubility at different pH-values to document the influence on the functional properties. Another objective of this paper was to demonstrate the influence on the digestibility of the proteins with one of the main enzymes of the gastro-intestinal tract (pancreatin) on the basis of in vitro experiments after derivatization with phenolic substances. The enzymatic digestion of the derivatized proteins was promoted.

Model studies on reactions of plant phenols with whey proteins.

Whey proteins were modified by reaction with selected phenolic compounds (ferulic-, chlorogenic-, caffeic- and gallic acid) and related substances (quinic acid and p-quinone) as well as with extracts from coffee, tea, potato and pear at pH 9. The derivatives formed were characterized in terms of their physicochemical and digestion properties. The derivatization was accompanied by a reaction at the lysine and tryptophan side chains, whereby their content was decreased in comparison to that in the control whey proteins. Moreover, the solubility of the derivatives decreased over a broad pH range and the derivatization influenced the hydrophobe-hydrophile character of the whey proteins. The isoelectric points were shifted to lower pH values in the order of reactivity as follows: gallic acid > p-quinone > caffeic acid > chlorogenic acid. The other derivatives showed no or few changes compared to the control whey proteins. The formation of high molecular fractions was documented with SDS-PAGE. Especially the derivatives of chlorogenic-, caffeic-, gallic acid and p-quinone showed an increase in molecular weight of beta-lactoglobulin fraction from 18,300 to 20,000 Da. A dimer formation in molecular range 40,000 was also registered. MALDI-TOF-MS was applied to characterize the binding of the individual phenolic compounds or their oxidation products to the whey protein fractions, alpha-lactalbumin and beta-lactoglobulin. In vitro experiments showed that the digestion of the derivatized whey proteins with the enzymes of the gastrointestinal tract (trypsin, chymotrypsin, pepsin and pancreatin) was adversely effected. Similar results with regard to physicochemical characterization and digestion properties of the whey proteins treated with the applied extracts from plant beverages, fruit and vegetable were also documented. Coffee and tee were comparatively the most reactive extracts.

Physicochemical properties and susceptibility to proteolytic digestion of myoglobin-phenol derivatives.

This paper deals with the interactions of chlorogenic, caffeic, and quinic acids and p-quinone with myoglobin. The myoglobin derivatives formed have been characterized in terms of physicochemical properties and susceptibility to proteolysis. The results show that the free amino group and tryptophan contents of the myoglobin-phenol derivatives decrease with the increasing extent to which the protein becomes derivatized. Furthermore, the solubility of myoglobin-phenol derivatives decreases in the pH range 3.5-6.5 as compared to solubility of the native protein. The reaction also influences the hydrophilic-hydrophobic character of the protein. The isoelectric point of the derivatized myoglobin is shifted to a lower pH value, and formation of high molecular fractions is also documented. This paper also demonstrates the influence of the protein derivatization with plant phenols on susceptibility to digestion by trypsin, alpha-chymotrypsin, and pepsin, determined in vitro. The enzymatic digestion of the derivatized proteins is adversely affected.

Reactions of a glucosinolate breakdown product (benzyl isothiocyanate) with myoglobin.

The interaction of various amounts of benzyl isothiocyanate (benzyl-ITC) with myoglobin is known to lead to the formation of derivatives. These have been characterised by the determination of solubility, free amino group, tryptophan content and chromatographic as well as electrophoretic behaviour. In the range between 2.5 and 125 mg benzyl-ITC/g protein, all properties of the reaction products correlate with the concentration of benzyl-ITC. However, at 250 mg benzyl-ITC/g myoglobin, a rather unexpected low degree of derivatization, as well as atypical chromatographic and electrophoretic behaviour, is observed. The proposed explanation was that conformational changes in the presence of a high concentration of hydrophobic benzyl-ITC made fewer amino groups accessible to the reagent. To test this hypothesis we have run the reaction under denaturing conditions. The results showed that the reaction of myoglobin with high concentrations of benzyl-ITC in the presence of 8 M urea led to a higher degree of derivatization than in the presence of water only. In addition, the Mr distribution of the reaction products was determined by MALDI-TOF-mass spectrometry and the overall degree of derivatization calculated from the spectra.

Some aspects of reactions of benzyl isothiocyanate with bovine sarcoplasmic proteins.

Benzyl-ITC (benzyl isothiocyanate) reacts preferentially with amino groups and sulfhydryl side chains of bovine sarcoplasmic proteins to form thiourea and dithiocarbamate derivatives, generally resulting in a decrease in solubility of the derivatives along a wide pH range. Under these conditions, it was also possible to show that secondary amine side chains, as found in tryptophan, also react with benzyl-ITC. A quenching of tryptophan fluorescence intensity after interaction with benzyl-ITC was also observed. Polyacrylamide gel electrophoresis (PAGE) experiments in the presence of urea indicated changes in electrophoretical mobility and in composition of subfractions. Changes in surface hydrophobicity and composition as determined by the ANS (8-anilinonaphthalene-1-sulfonate) method and RP-HPLC were also observed. Polymerisation of protein molecules after reaction with benzyl-ITC was documented using the SDS-PAGE technique. These investigations showed that a group of subfractions belonging mainly to glycolytic enzymes and associated proteins with molecular weight between 38-70 kDa was highly reactive.

Selected physico-chemical properties of succinylated legumin from pea (Pisum sativum L.).

Selected physico-chemical properties of pea legumin before and after succinylation have been investigated using isoelectric focusing, PAGE, SDS-PAGE, hydrophobicity measurements, SE-HPLC and RP-HPLC. Exhaustive succinylation shifted the I.P. of legumin from 4.75 to 3.5. The stepwise dissociation of legumin by increasing succinylation has been confirmed both by means of PAGE in a nondenaturing system, and by SE-HPLC. The results of SDS-PAGE provided evidence for the exposure of alpha-polypeptide chains in the native legumin. High succinylation resulted in a decrease of the surface hydrophobicity (S0) measured by both fluorescence probes (cis-parinaric acid and anilino-naphthalene sulfonic acid). RP-HPLC gave a response both to conformational changes and the introduced succinyl residues.