4 . 2013
Biochemistry of nutrition

Polymorphisms Ser447Ter of lipoprotein-lipase gene, Cys112Arg and Arg158Cys of apolipoprotein E gene in patients with obesity

The distribution of allele Ser447Ter of lipoprotein lipase gene (LPL) and polymorphic markers E2 and E4 of the apolipoprotein E gene (ApoE) were examined in 100 obese patients at the age of 18–66 years (28 men and 72 women, 40,6±2,1 years old). The first group included patients with I degree of obesity (n=26, BMI=32,5±0,2), the second group – patients with II degree of obesity (n=33, BMI=37,1±0,2), the third group – patients with grade III obesity (n=41, BMI=46,3±1,1) and control group were 18 healthy individuals aged from 22 to 55 years (7 men and 11 women, 36,5±0,9 years old, BMI=22,4±1,8). Maximal frequency of allelic polymorphism ε2 has been revealed in patients with I degree of obesity, and allele ε4 – in patients with III degree of obesity. The most common genotype of ApoE gene was ε3/ε3 in all three groups of patients with obesity. In a comparative analysis of allelic variants of the Apo E gene occurrence it has been found that the frequency of a polymorphic variant ε2/ε2 tended to decrease with BMI increasing, whereas a higher rate of detection of genotypes ε4/ε3, ε4/ε4 and ε2/ε4 was found in patients with III degree of obesity. The data obtained suggest that the ε4 allele of the Apo E gene is associated with the development of morbid obesity, rather than allele ε2. This phenomenon can be explained by the fact that apoE4 isoform has reduced affinity for LDL in comparison with apolipoprotein E3. The maximum concentration of cholesterol, triglycerides and LDL cholesterol has been observed in patients with ε2/ε4 genotype of ApoE gene, and it was significantly higher than in the control group (p<0,05). The content of blood lipid fractions in patients with ε3/ε4 genotype of ApoE gene, in contrast, was the lowest among obese and did not exceed the values of the control group (p>0,05). These data indicates a small contribution of ε4 polymorphism in heterozygous form to the development of dyslipidemia in obesity. The most positive effect of diet treatment was achieved in patients with genotype ε3/ε3 and ε3/ε4. An integrated approach to the assessment of lipid metabolism in patients with obesity, including the analysis of polymorphic genetic loci, can optimize and personalize the diet therapy.

Body structure and serum concentration of adiponectin and cytokines (IL-6, -10 and TNF-α) in rheumatoid arthritis combined with obesity

In the study the comparative assessment of the main markers of body structure and serum concentrations of adiponectin and cytokines (IL-6, TNF-α, IL-10) has been carried out in 26 rheumatoid arthritis patients at a normal body mass index (BMI) [Ме; 25; 75 percentile = 24,1 (23,5; 28,6)] and in 15 obese patients [BMI = 33,4 (31,3; 42,7)]. The control group consisted of 25 apparently healthy individuals [BMI = 23,5 (21,8; 24,4)] randomized by sex and age with the main group. The expressed increase of production of pro-inflammatory cytokines – TNF-α and IL-6 in both groups has been established, in the group of patients with obesity serum concentration of these cytokines 1,8–2,4 fold exceeded similar in group with a normal BMI and amounted 245,3 (89,3; 302,3) and 54,4 (18,4; 72,3) pg/ml correspondingly. Anti-inflammatory cytokine IL10 increased more significantly at normal BMI [227,2 (143,4; 282,3) pg/ml)] in comparison with group of rheumatoid arthritis patients with obesity [122,2 (89,3; 164,3) pg/ml] while in control group its content was 60,8 (24,3; 75,4) pg/ml. It has been shown that serum concentration of adiponectin was higher in group with normal BMI [9,2 (6,3; 15,3) mcg/ml] and was lower in patients with obesity [3,2 (2,3; 8,3) mcg/ml] than in healthy subjects [7,4 (4,4; 9,2) mcg/ml]. The analysis of the body structure markers demonstrate that fatty tissue content increase both in patients with normal BMI (1,7 fold) and at obesity (2,6 fold).

Hematological indices of rats having complete and vitamin-deficient diets enriched with dietary fibers

The hematological indices in 48 rats Wistar male with initial body weight 58,1±0,5g has been studied. The rats were divided into 6 group and fed the complete semi-synthetic diet, containing 100% or 20% of vitamin mixture (Vit) with or without addition of dietary fiber (DF) in the form of wheat bran (5% of diet mass) during 4 weeks. The animals of the 1 group received 100% of vitamin mixture (100% Vit); of the 2 group – 100% Vit+DF; 3 group – 20% of vitamin mixture (20% Vit); 4 group – 20% of vitamin mixture and DF (20% Vit+DF). The next 5 days rats from vitamin-deficient groups were fed with diets supplemented with 80% of Vit: (5 group – 20% Vit+ 80% Vit; 6 group – 20% Vit+DF+80% Vit). The animals fed vitamin-deficient diet lag significantly in growth from animals fed a complete diet. The growth curve of rats fed a diet with DF took an intermediate position. The studies were carried out at the Hematology analyzer «Coulter AC TTM 5 diff OV» (Beckman Coulter, USA) under the program, developed for the study of rat blood. Mean corpuscular volume (MCV) and mean corpuscular hemoglobin content (MCHC) were significantly decreased in rats with combined vitamin deficiency (20% Vit) in the diet compared with those of control group rats, while the compensatory increase in the number of red blood cells (RBC) and leukocytopenia took place. The enrichment of vitamin-deficient diet with DF (20% Vit+DF) prevented MCV and RBC changes, but MCHC left reduced in comparison with the indicator of the rats in control group. Indicators characterizing the state of platelets had no statistically significant differences between the groups. Compensation of vitamin deficiency in the diet of rats from group 5 (20% Vit+80% Vit) led only to the normalization of MCV. After vitamin restore in the diet of group 6 (20% Vit+DF+80% Vit) all investigated parameters were indistinguishable from the parameters of the control group. This indicates a positive effect of wheat bran DF consumption on the studied hematological indices.

Effect of intragastrically injection of phytoecdysteroids on some indicators of hormonal status in rats

The experiment in vivo in growing male Wistar rats was carried out. The animals of the experimental 2–4 groups were daily intragastrically injected water solutions of the dried extract from the leaves of Seratulla coronata L. in volume of 1,0 ml, containing 2, 20 and 50 mg of phytoecdysteroids per kg of animal weight, accordingly. Animals of control group were daily injected 1,0 ml of water. The content of phytoecdysteroids in the dry extract was analyzed by high performance liquid chromatography (HPLC). The concentration of the sum of phytoecdysteroids in dry extract was 6,15%, 66% of which was 20-hydroxyecdysone and 23% was 25S-inokosteron. On the 15th day animals were taken out of the experiment by the decapitation. The content of corticosterone, prostaglandin E2 and beta-endorphin in rat blood plasma were determined by ELISA test. The pathological – anatomical analysis and weighing of the liver did not reveal any adverse changes of this organ in the animals of all groups. The average concentration of blood plasma corticosterone reduced with increasing of the dose of the extract injected to the animals, reaching significant differences relative to the control group (60,9±9,4 ng/ml) for 3 and 4 groups (22,7±6,6 and 17,6±7,3 ng/ml, accordingly). Beta-endorphin and prostaglandin E2 levels did not differ. The ratio of the mediator of stress (corticosterone) and inhibitors of stress (beta-endorphin and prostaglandin E2) has been calculated. A monotonic decrease of corticosterone/beta-endorphin and corticosterone/prostaglandin E2 ratio has been found with extract dose increasing. Taken together the results of determination of biochemical parameters of the general adaptation syndrome the dose-dependent stress-protective effect of Seratulla coronata L extract has been demonstrated.

The state of the antioxidant status of children of different ages

Тhe parameters of lipid peroxidation (blood plasma content of primary products of lipid peroxidation and the end of TBA-active products) and indexes of antioxidant protection (total antioxidant activity, the content of fat-soluble vitamins A and E in blood plasma and riboflavin in blood) has been evaluated in 75 healthy children living in Irkutsk. All respondents were divided into three age periods: preschool age (3–6 years old, mean 4,7±1,0 – 21 children), primary school age (7–8 years, 7,6±0,4, – 28 children) and middle-school age (9–11 years, 9,9±0,7, 26 children). We used spectrophotometric and fluorometric methods. Identified the distinctive features of the metabolic reactions in children at different ages. The increase of the content of the primary products of lipid peroxidation in the early school years and of the end of TBA-active products for children of secondary school age in comparison with the preschool children has been determined. At the same time an increase in the level of total antioxidant activity and fat-soluble vitamins and riboflavin content in young and middleschool aged children in comparison with the pre-school children has been observed. The measurement of vitamins status demonstrated the insufficiency of α-tocopherol in half of pre-school children, in 36% of primary school children and 38% of children of secondary school age. Retinol and riboflavin deficiency has been detected in a small proportion of children of all ages. In this regard, the additional supply with vitamins of children in pre-school and middle-school period is essential.

Allergic reactions to cereals in children with atopy

Cereals are the necessary and the major products in the diet. The aim of the study was to determine the dependence of frequency of sensitization to various cereals in children with atopy upon the age and diagnosis on the basis of anamnesis, the results of elimination diets, skin tests and the presence of specific IgE antibodies. 416 children and adolescents aged from 6 months to 17 years with atopic dermatitis (n=359) and asthma (n=57) has been investigated. The frequency of sensitization to cereals in children with atopic dermatitis according to skin tests ranged from 31 to 43%. Expressed sensitization (skin test +++, ++++) on the large majority of groats did not exceed 3% and only on corn and barley, it took place in 5,8 and 3,8%, respectively, while 81% of these children data coincided with a case history. The percentage of severe sensitization in boys was generally higher than that among girls. The percentage of pronounced sensitization to cereals increased with age in patients with atopy. The reason why the frequency of marked sensitization to maize in asthmatic children older than 7 years reached 20% and in children with atopic dermatitis reached 7,5% at the age of younger 1 year and 7,7% at the age from 3 to 7 years, remains unclear and requires further study.

Correction of polyhypovitaminosis in rats, having standard and enriched with polyunsaturated fatty acids family ω-3 diets with different doses of vitamins

The efficiency of the correction of combined alimentary vitamin deficit in male Wistar rats (body weight 90–121 g) fed standard diet or enriched with ω-3 polyunsaturated fatty acids diet (by replacing sunflower oil (4,5% of the diet) with linseed oil) with different doses (physiological and enhanced) of vitamins has been investigated. The control group of animals (n=12) received a complete semisynthetic diet during all experimental period (42 d). The animals of the test groups (each of 20 rats) received vitamin-deficient diet containing sunflower or linseed oil and 20% per cent of vitamin mixture amount in control diet from which vitamin E had been excluded. After 28 days of such feeding each of test groups was divided into two subgroups (6–8 rats in each), and the next 14 days the animals of subgroups received the diet with different degree of vitamin compensation (50 or 180% percent of vitamin content in the diet of the control group). The addition of both low and high vitamin dose in deficient diet based on standard fat component did not compensate the reduced liver vitamin A content, which amounted to 47,4% of the level in the liver of the control group. The lack of vitamin E in animals was eliminated only after adding of the enhanced dose of vitamin E to the ration. Recovering of decreased plasma and liver B2 level, plasma 25(ОН)D and liver vitamin B1 content have been occurred after addition of the low dose of these vitamins to rat diet. Increasing of ω-3 PUFA diet level improved vitamins A and D sufficiency to some extent, but was accompanied by the significant reduction of rat liver alfa-tocopherol content both under combined vitamin deficiency (by 14%) and increased vitamins consumption (by 43%). PUFA enrichment of the diet of rats with vitamin deficiency had no impact on vitamin B1 and B2 liver level. The use of high doses of vitamins for a long time to eliminate a combined deficiency of vitamins has been proved.
Research methods

Direct detection of T-2- and HT-2-Mycotoxins producers of fungi the genus Fusarium in food grain by PCR (report 2)

The improvement of the Fusarium DNA extraction method has been undertaken in order to reduce the error of PCR analysis for detection of toxigenic Fusarium species, including those contained in the grain in the uncultureted state, directly in the grain. The efficiency of Fusarium DNA extraction methods (nucleotides sorption and CTAB method) has been compared. The efficiency of CTAB method combined with 10-fold weight increase of milled grain sample has been demonstrated. This approach revealed a greater number of Fusarium species, than PCR analysis of combined Fusarium mycelium from the same samples. The uncultureted F. langsethiae was detected in the DNA extract from a sample of barley, which was not identified in the combined sample of the mycelium. This sample of the grain has the highest levels of T-2/NT-2toxins – 0,075/0,345 mg/kg (determined by HPLC) among positive samples. F. sporotrichioides – a potential producer of T-2- and HT-2-toxins has been revealed by PCR method in other grain samples both containing and not containing these toxins The biosynthesis of T-2- and HT-2-toxins on the PSAmedium in vitro has been studied for 10 single-spores F. sporotrichioides isolates, allocated from grain. Synthesized T-2-toxin content (measured by ELISA) ranged from 0,4 to 184,5 mg per l of medium. Three strains showed very high levels from 117,2 to 184,4 mg/l, two of which have been isolated from barley which don’t contain these toxins. The absence of the toxin in grain samples does not guarantee the absence of high-level producers of mycotoxins. The direct detection of Fusarium spp. in grain by PCR analysis with extraction of fungal DNA by CTAB method along with increased sample weight has been shown to make possible the detection of a more number of species of Fusarium (including uncultureol strains) compared with mycological method with PCR analysis of the combined sample of the mycelium.
Diet treatment

Influence of dietotherapy enriched with conjugated linoleic acid on anthropometrical indicators and body composite structure in patients with an overweight

The clinical efficiency of dietotherapy enriched with conjugated linoleic acid (3 g per day) in patients, suffering from the excess mass of a body and obesity of I and II degrees has been investigated. Dynamics of anthropometrical indicators and body composite structure before and after the treatment of patients from the main (n=20) and control groups were estimated. Studied parameters were fixed in day 0, 14 and 28. Statistically significant reduction of the weight, body mass index (BMI), anthropometrical parameters (volumes of a waist, hips, a shoulder), indicators of body composite structure (fatty weight, area of splanchnic fat, muscular weight) has been revealed as a result of the carried-out course of treatment, and they were more expressed in patients of the main group. In particular, the waist size authentically decreased by 6,5% in patients from the main group, while in patients from the group of comparison the decrease reached only 4,2%. Average change of waist volume was – 7,0 cm in the main group and – 4,5 cm in the control group. The grasp of hips in the main group decreased by 4,2%, in the group of comparison for 2,8%. Absolute change of hips volume in the main group 1,67 fold exceeded that in the group of comparison (р<0,005). The shin grasp in the main group decreased by 9,7%, while in the control group remained invariable. BMI decreased by 7,2% and 5%, respectively. Thus, the reduction of body fatty weight was 10% in patients from the main group and 4% in patients from the group of comparison. The body total liquid decreased by 1,4 l in the main group, and only by 0,3 l in the group of comparison. The received intergroup distinctions are statistically reliable (р<0,05).

Legislative regulation of production and turnover of products for people with different diseases

This article presents results of analysis of existing regulatory documents and approaches to the legislative regulation of production and turnover of special dietary products for people with specific diseases in EU, Ukraine and Russian Federation. According to the EU legislation, production and turnover of food products for nutritional support of people during specific diseases and the rehabilitation period are regulated by the Commission Directive 1999/21/EС, 2009/39/ES, by Regulation Commission (EU) № 953/2009 and documents of Codex Committee. Special food products for people with specific diseases in Ukrainian legislation are classified as nutrition products for special dietary use and are regulated by the following Laws of Ukraine «On the safety and quality of food», «On ensuring of sanitary and epidemiological welfare of the population», «On Consumer Rights Protection», «On advertising» and by other non-legislative acts. According to the current legislation of the Russian Federation, the products for people with specific diseases are classified as healthy dietary food products. The basis of the legal framework are federal laws «On the quality and safety of food», «On the sanitary-epidemiological welfare of the population», «On technical regulation» and technical regulations of the Customs Union «On Food Safety» and «On the safety of certain types of specialized food products, including healthy dietary food products and therapeutic dietary food products». There is no common approach to the legal regulation of production and turnover of products for people with specific diseases in the world. The proposals for further harmonization of regulatory control in this area have been developed.
Nutrition of sportsmеn

Investigation of actual nutrition and some factors of physical status in rugby players

Physical status and nutrition and energy value of factual diets of 18 rugby players (15–30 years old) assessed during current medical observation were analyzed. Anthropometric data (height, body mass – BM, waist circumference/hip circumference index, body mass index – BMI), body composition data and resting metabolic rate by bioimpedance method (analyzer ABC-01 «Medass») were determined. These indices were found to be normal except more high level of BMI due to high levels of lean and skeletal muscle mass, but not a result of obesity. Estimation of factual nutrition based on 1-day feeding (nutritionscreening questionnaire method) has demonstrate that it was not optimal both for energy and nutrition value for this group of physical activity. Energy value ranged from 1889 kcal to 3175 kcal (22,5–35,1 kcal/kg BM/day) with lower carbohydrate intake (46,9%) and higher fat intake (35,8%) were found. Nutrition value ranges: protein – 67–144 g (0,71–1,38 kcal/kg BM/day), fat – 75–140 g (0,88–1,51 kcal/kg BM/day), carbohydrate – 173–420 g (2,17–4,66 kcal/kg BМ/day) were observed. The findings suggest the need to pay more attention to in-depth study of dietary intake of athletes to maintain health and to create optimal conditions for a professional sports activities.

Vitamin and mineral status of oarsmen during the training-competition cycles

Vitamin and minerals status of 30 sportsmen aged 16,8±0,2 years (trained in boat racing 5,0±0,3 years) during the training – competition cycles in autumn and winter has been studied. A significant decrease of vitamin A level in the range of reference limits was found in 30,8% of sportsmen during autumn season, but such decrease was revealed in 100% of examined sportsmen during winter. Vitamin B1 and B2 deficiency has been detected in both seasons.The portion of persons with vitamin B2 deficiency increased during training cycles. A significant decrease of magnesium (in 56,3% of sportsmen) and sodium level (in 33,3% of sportsmen) was detected during autumn season. Irrespective of the season a significant decrease of blood serum of calcium (in 40,0–66,7% of sportsmen); potassium (in 53,3–66,7% of sportsmen according to both absolute and individual data); chlorides (in 50,0–90,0% of sportsmen), and iron (in 53,3–60,0% of sportsmen) has been revealed. The data obtained indicate the need for diagnostics and correction of vitamin and minerals status in sportsmen.
Hygiene of nutrition

Food contamination by pathogens of acute intestinal infections and survival of microorganisms in them under some variants of thermal processing and storage

The contamination of the surface of vegetables and fruits with conditionally pathogenic enterobacteria, intestinal protozoa and enteric viruses has been revealed. The experiment showed an opportunity of penetration of enteric viruses inside of vegetables and fruits with intact (without visible damages) surface. S. enteritidis survive on the shells of the eggs 4 months in the refrigerator, in the albumen and yolk – more than 5 months and in the long storage (more than one month) of chicken eggs in the refrigerator S. enteritidis could penetrate into the egg through the intact shell and multiply in the yolk. It was shown in the experiment on the condition laying of chicken eggs of room temperature in cold water and their subsequent cooking S. enteritidis full destruction of pathogens comes on the 5 minute of heat processing, when the temperature inside the eggs reaches more than 80 °С. In mayonnaise at the temperature of the refrigerator S. еnteritidis reproduce, but remain viable for 13 days. In the egg powder at the room temperature noted smooth death of S. enteritidis, term viability of microorganisms is 5 months. Addition 10% acetic acid to egg powder in the ratio 1:2 provides for the death of S. enteritidis for a minute. In experimental conditions was established that the processing of chicken on the grill in the process of cooking «Shaverma» provides exemption meat from Salmonella only in the thickness of a piece of up to 2 centimeters. The deeper layers of meat and meat juice which accumulates on the pallet grill, throughout the duration of thermal processing may remain contaminated Salmonella.

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Viktor A. Tutelyan
Full Member of the Russian Academy of Sciences, Doctor of Medical Sciences, Professor, Scientific Director of the Federal Research Centre of Nutrition, Biotechnology and Food Safety (Moscow, Russia)

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