Effect of plant polyphenols on the formation of advanced glycation end products from ß-lactoglobulin.

Dietary exposure to advanced glycation end products (AGEs) formed from proteins and reducing sugars is of increasing concern to human health. AGEs may form in protein-based powders containing sugars for instant beverages during drying and storage of the product. Chlorogenic acid, a plant phenol characteristic of coffee, was found to protect against the formation of AGEs at a concentration of 50mM during heating of ß-lactoglobulin in the presence of glucose as a reducing sugar in 30% aqueous ethanol at 70°C. Epicatechin, a plant phenol characteristic of green tea, had no similar effect for the equivalent concentration of phenol on the formation of AGEs. Immunochemical detection (ELISA) using polyclonal antibodies raised against AGEs showed a dose-dependent effect of protection by chlorogenic acid on AGE formation and is recommended for routine quality control of sugar containing milk-based powders for instant beverages.

Short-term effects of dietary advanced glycation end products in rats.

Dietary advanced glycation end products (AGE) formed during heating of food have gained interest as potential nutritional toxins with adverse effects on inflammation and glucose metabolism. In the present study, we investigated the short-term effects of high and low molecular weight (HMW and LMW) dietary AGE on insulin sensitivity, expression of the receptor for AGE (RAGE), the AGE receptor 1 (AGER1) and TNF-α, F2-isoprostaglandins, body composition and food intake. For 2 weeks, thirty-six Sprague-Dawley rats were fed a diet containing 20% milk powder with different proportions of this being given as heated milk powder (0, 40 or 100%), either native (HMW) or hydrolysed (LMW). Gene expression of RAGE and AGER1 in whole blood increased in the group receiving a high AGE LMW diet, which also had the highest urinary excretion of the AGE, methylglyoxal-derived hydroimidazolone 1 (MG-H1). Urinary excretion of N ε-carboxymethyl-lysine increased with increasing proportion of heat-treated milk powder in the HMW and LMW diets but was unrelated to gene expression. There was no difference in insulin sensitivity, F2-isoprostaglandins, food intake, water intake, body weight or body composition between the groups. In conclusion, RAGE and AGER1 expression can be influenced by a high AGE diet after only 2 weeks in proportion to MG-H1 excretion. No other short-term effects were observed.

Quantification of radicals formed during heating of ß-lactoglobulin with glucose in aqueous ethanol.

Heating ß-lactoglobulin alone, or in the presence of glucose, at moderately elevated temperatures (50, 70 and 80°C) in 30% aqueous ethanol at pH 6.0, 7.0 and 8.0 led to an iron catalysed formation of highly reactive radicals at concentrations of up to 1.0 mmol/mol protein as trapped by α-(4-pyridyl N-oxide)-N-tert-butylnitrone (POBN) and quantified by ESR spectroscopy. Oxidation of ß-lactoglobulin was favoured by increasing temperature and high pH conditions, whereas presence of glucose decreased oxidation, indicating that oxidation of ß-lactoglobulin involves either amine or thiol side chain group blocked as oxidation substrate by reaction with glucose as a reducing sugar. The oxidation of ß-lactoglobulin was hampered by the phenolic antioxidant 4-methylcatechol and to an even larger extent by the oxidised quinone form 4-methyl-1,2-benzoquinone, suggesting that it is the quinone forms of phenolic antioxidants which compete with reducing sugars in protecting proteins against oxidation.

Oxidative stability and chemical safety of mayonnaise enriched with grape seed extract.

Grape seed extract (GSE), a by-product from the wine industry, was explored for its use as enrichment to mayonnaise, due to its potential health effects. Mayonnaises were enriched with 0 mg GSE per mL, 0.5 mg GSE per mL (~0.050%), 0.9 mg GSE per mL (~0.10%) and 1.4 mg GSE per mL (~0.15%) during preparation and stored in the dark at room temperature for 8 weeks. The antioxidative capacity of the mayonnaises was evaluated by their ability to scavenge the stable radical TEMPO by electron spin resonance (ESR) spectroscopy. The oxidative stability of the mayonnaises was determined by the content of lipid hydroperoxides (peroxide value, POV), the content of conjugated diene hydroperoxides and the content of thiobarbituric acid reactive substances (TBARS). The highest antioxidative capacity and the lowest content of lipid hydroperoxides and TBARS were found in the mayonnaise with the highest percentage of GSE (0.15%). Therefore, the oxidative stability of the mayonnaises enriched with GSE was slightly improved through storage. However, mayonnaise without GSE had the highest sensorial acceptability compared to mayonnaise enriched with GSE. In the Artemia salina assay, a fast screening method for overall toxicity, the death rate of brine shrimps larvae was found to increase for increasing percentage of GSE. A level of 0.05% GSE in mayonnaise is concluded not to constitute any toxicological risks, but to provide significant protection against oxidation during storage.

Palatability and chemical safety of apple juice fortified with pomegranate peel extract.

Pomegranate peel extract (PPE), a by-product of the pomegranate juice industry with potential health effects, was explored for use to fortify reconstituted apple juice in the concentration range 0.5 to 2.0% (w/w). Radical scavenging and antioxidative capacities of the fortified apple juices were evaluated using (i) electron spin resonance (ESR) to quantify their ability to scavenge the stable radical Fremy's salt and (ii) the Trolox equivalent antioxidant capacity (TEAC) assay and compared to apple juice without fortification as control. The highest antioxidative capacity was found in the apple juice fortified with the highest percentage of pomegranate peel extract, while the optimal sensory quality was found by addition of 0.5 g PPE per 100 mL. The Artemia salina assay was used as a fast screening method for evaluating overall toxicity, and showed little toxicity with up to 1.0 g per 100 mL addition of PPE, but increasing toxicity at higher concentrations. Accordingly, it is important to balance addition of PPE, when used for enrichment of apple juice in order to obtain a healthier product, without compromising the sensorial quality or toxicological safety of the apple juice. Concentrations between 0.5 and 1.0 g PPE per 100 mL seem to be acceptable.

Advanced glycation endproducts in food and their effects on health.

Advanced glycation endproducts (AGEs) form by Maillard-reactions after initial binding of aldehydes with amines or amides in heated foods or in living organisms. The mechanisms of formation may include ionic as well as oxidative and radical pathways. The reactions may proceed within proteins to form high-molecular weight (HMW) AGEs or among small molecules to form low-molecular weight (LMW) AGEs. All free amino acids form AGEs, but lysine or arginine side chains dominate AGE formation within proteins. The analysis of AGEs in foods and body fluids is most often performed by ELISA or LC-MS; however, none of the methodologies cover all HMW and LMW AGEs. Most research is, therefore, carried out using 'representative' AGE compounds, most often N(ε)-carboxymethyl-lysine (CML). Only LMW AGEs, including peptide-bound forms, and carbonyls may be absorbed from the gut and contribute to the body burden of AGEs. Some AGEs interact with specific pro- or anti-inflammatory receptors. Most studies on the biological effects of AGEs have been carried out by administering heated foods. The pro-inflammatory and deteriorating biological effects of AGEs in these studies, therefore, need further confirmation. The current review points out several research needs in order to address important questions on AGEs in foods and health.

Total and inorganic arsenic in dietary supplements based on herbs, other botanicals and algae--a possible contributor to inorganic arsenic exposure.

The content of total and inorganic arsenic was determined in 16 dietary supplements based on herbs, other botanicals and algae purchased on the Danish market. The dietary supplements originated from various regions, including Asia, Europe and USA. The contents of total and inorganic arsenic was determined by inductively coupled plasma mass spectrometry (ICP-MS) and anion exchange HPLC-ICP-MS, respectively, were in the range of 0.58 to 5.0 mgkg(-1) and 0.03 to 3.2 mg kg(-1), respectively, with a ratio between inorganic arsenic and total arsenic ranging between 5 and 100%. Consumption of the recommended dose of the individual dietary supplement would lead to an exposure to inorganic arsenic within the range of 0.07 to 13 µg day(-1). Such exposure from dietary supplements would in worst case constitute 62.4% of the range of benchmark dose lower confidence limit values (BMDL01 at 0.3 to 8 µg kg bw(-1) kg(-1) day(-1)) put down by European Food Safety Authority (EFSA) in 2009, for cancers of the lung, skin and bladder, as well as skin lesions. Hence, the results demonstrate that consumption of certain dietary supplements could contribute significantly to the dietary exposure to inorganic arsenic at levels close to the toxicological limits established by EFSA.

Antioxidant synergism between ethanolic Centella asiatica extracts and α-tocopherol in model systems.

The synergistic antioxidant effects of ethanolic extracts of Centella asiatica (CE), and α-tocopherol have been studied. The types of interactions exhibited by CE and α-tocopherol combined at different ratios were measured using three assays: 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt (ABTS) radical-scavenging capacity, the ß-carotene bleaching system and liposome peroxidation assays. Fixed-fraction isobolographic analysis was used to detect any inducement of the antioxidant activity compared with the individual activities of CE and α-tocopherol. Of all synergistic combinations of CE and α-tocopherol, only fraction 2/3 showed the synergistic combination that fits well in three different assays and can be explained by the regeneration of α-tocopherol by CE despite the interaction effect of ß-carotene present in the analytical assay. This phenomenon involved complex interactions between CE and α-tocopherol to exhibit different degrees of interactions that eventually increased antioxidant activity.

Antioxidant capacity versus chemical safety of wheat bread enriched with pomegranate peel powder.

Pomegranate peel powder (PP), a by-product of the pomegranate juice industry rich in polyphenols, was explored for use in bread production, due to its potential health effects. Wheat bread was prepared using different levels for replacement of flour with PP (0 to 10 g per 100 g flour) resulting in antioxidant levels expressed as Trolox equivalent antioxidant capacity values (TEAC) ranging from 1.8 to 6.8 µmol TEAC per g bread for fresh bread. TEAC remained constant during 5 days of storage in polyethylene bags at room temperature. The oxidative stability was evaluated by detection of radicals by direct electron spin resonance (ESR) spectroscopy, and peroxide value, and the highest capacity of scavenging of radicals (Fremy's salt) and the lowest content of peroxide values were found in bread with the highest percentage of PP. Safety evaluation was performed by the Artemia salina assay. An increased death rate of the brine shrimp larvae was found as a function of the replacement of wheat flour with PP in fortified bread providing a general screening method for the toxicological test of polyphenol fortified bread to be recommended for use in product development in addition to subjective evaluation. Based on both toxicological and subjective evaluations an addition of 2.5% PP is recommended for the actual product.

Occurrence and sorption properties of arsenicals in marine sediments.

The content of total arsenic, the inorganic forms: arsenite (As(III)) and arsenate (As(V)), the methylated forms: monomethylarsonic acid and dimethylarsinic acid (DMA), trimethylarsenic oxide, tetramethylarsenonium ion and arsenobetaine was measured in 95 sediment samples and 11 pore water samples from the Baltic Sea near the island of Bornholm at depths of up to 100 m. As(III+V) and DMA were detected in the sediment and As(III+V) was detected in the sediment pore water. Average total As concentration of 10.6 ± 7.4 mg/kg dry matter (DM) in the sediment corresponds to previously reported values in the Baltic Sea and other parts of the world. Existing data for on-site measurements of sorption coefficients (Kd) of arsenicals in marine and freshwater sediments show large variability from <1 to >1,000 L/kg. In this work, calculated sorption coefficients (Kd and Koc) for As(III+V) showed significant correlation with depth, dissolved oxygen (DO), salinity and sediment classification; for depths <70 m, salinity <11 %, DO >9 mg/L and sand/silt/clay sediments the Kd was 118 ± 76 L/kg DM and for depths >70 m, salinity >11 %, DO < 9 mg/L and muddy sediments the Kd was 513 ± 233 L/kg DM. The authors recommend using the found Kd value for arsenic in marine sediments when conditions are similar to the Baltic Sea. At locations with significant anthropogenic point sources or where the local geology contains volcanic rock and sulphide mineral deposits, there may be significantly elevated arsenic concentrations, and it is recommended to determine on-site Kd values.

Development and validation of an SPE HG-AAS method for determination of inorganic arsenic in samples of marine origin.

The present paper describes a novel method for the quantitative determination of inorganic arsenic (iAs) in food and feed of marine origin. The samples were subjected to microwave-assisted extraction using diluted hydrochloric acid and hydrogen peroxide, which solubilised the analytes and oxidised arsenite (As(III)) to arsenate (As(V)). Subsequently, a pH buffering of the sample extract at pH 6 enabled selective elution of As(V) from a strong anion exchange solid-phase extraction (SPE) cartridge. Hydride generation atomic absorption spectrometry (HG-AAS) was applied to quantify the concentration of iAs (sum of As(III) and As(V)) as the total arsenic (As) in the SPE eluate. The results of the in-house validation showed that mean recoveries of 101-104% were achieved for samples spiked with iAs at 0.5, 1.0 and 1.5 mg·kg(-1), respectively. The limit of detection was 0.08 mg kg(-1), and the repeatability (RSD(r)) and intra-laboratory reproducibility (RSD(IR)) were less than 8% and 13%, respectively, for samples containing 0.2 to 1.5 mg kg(-1) iAs. The trueness of the SPE HG-AAS method was verified by confirming results obtained by parallel analysis using high-performance liquid chromatography coupled to inductively coupled plasma mass spectrometry. It was demonstrated that the two sets of results were not significantly different (P < 0.05). The SPE HG-AAS method was applied to 20 marine food and feed samples, and concentrations of up to 0.14 mg kg(-1) of iAs were detected.

Acrylamide-asparagine relationship in baked/toasted wheat and rye breads.

Acrylamide in baked and toasted wheat and rye bread was studied in relation to levels of asparagine in flour, dough, bread and toasts. Asparagine was consumed during bread preparation resulting in reduced acrylamide content in the products. In wheat bread, 12% of the asparagine initially present in the flour (0.14 g kg(-1)) remained after yeast fermentation and baking; for rye bread, 82% of asparagine remained after sourdough fermentation and baking. Asparagine present in untoasted wheat bread had totally reacted after hard toasting. Toasted wheat and rye bread slices contained 11-161 and 27-205 microg kg(-1) acrylamide, respectively, compared to untoasted wheat and rye bread with <5 and 7-23 microg kg(-1) acrylamide, respectively. The dietary intake of acrylamide from bread (untoasted) of 2 microg day(-1) is relatively low; however, acrylamide exposure from bread increases several fold for people eating toasted bread.

Kinetics of formation of acrylamide and Schiff base intermediates from asparagine and glucose.

From the concentration of glucose and asparagine as reactants and of acrylamide as product each determined by LC-MS during reaction in an acetonitrile/water (68:32) model system at pH 7.6 (0.04M phosphate buffer) and from the relative concentration of the Schiff base intermediate, the decarboxylated Schiff base intermediate, the Amadori product and aminopropionamide determined in the same reaction mixtures at 120°C, 140°C, 160°C and 180°C for up to 16min, the energy of activation for formation of the Schiff base intermediate was found to have the value 50±2kJmol(-1), while the apparent activation energy for formation of acrylamide was 64.4±0.6kJmol(-1), for formation of the decarboxylated Schiff base intermediate 92±2kJmol(-1), and for formation of the Amadori compound 59±4kJmol(-1), respectively. At high temperature conditions, formation of the Schiff base is accordingly rate determining, while at lower temperatures, decarboxylation becomes rate determining. Aminopropionamide was only detected at reaction times at which acrylamide formation already is significant in favor of, a reaction path including direct formation of acrylamide from the decarboxylated Schiff base, rather than including dissociation of ammonia from aminopropionamide.

Oxidative stability of buttermilk as influenced by the fatty acid composition of cows' milk manipulated by diet.

Milk from cows fed a low-fat diet high in cereals designed to stimulate fat synthesis de novo was lower in unsaturated fatty acids (21.3%) than milk from cows fed a diet high in fat, mainly from roasted soy beans (41.3% unsaturated fatty acids). Buttermilk from the more unsaturated milk was less oxidatively stable during storage (at 4 degrees C, followed for 11 d) than buttermilk from the more saturated milk, as monitored both by primary lipid oxidation products (lipid hydroperoxides) and by the secondary lipid oxidation product, hexanal. Fat-soluble antioxidants, beta-carotene and alpha-tocopherol, analysed by HPLC, were not consumed during storage for either of the two types of buttermilk. In contrast, the antioxidative capacity of the serum phase decreased during storage as evaluated in a radical scavenging assay based on the semi-stable water-soluble radical nitrosodisulphonate (Fremy's salt). The time course for the decrease in water-soluble antioxidants was very similar for the two types of buttermilk suggesting that oxidation is initiated in the serum phase independently of fatty acid composition.