History of nutrition and acid-base physiology

Summary In the 17^th century the notion of nutrition and diet changed in northern European countries. First chemical experiments fostered the idea that salts resulted from a union of acids and bases. Digestion was no more regarded as a process of cooking but a succession of fermentations controlled by a balanced production of acids and alkali. Life seemed to depend on the equilibrium of acids and alkalis. In the 19^th century food was systematically analysed for the content of energy and macronutrients and first scientifically based nutritional standards were formulated. The preferred use of processed food from the new food industry resulted in epidemics of nutritional disorders. Acidosis seemed to be a plausible pathogenic factor. Practitioners (S Ishizuka, H Hay, FX Mayr) formulated holistic doctrines integrating the concept of balance of acids and bases and recommending food with an excess of alkali. New micromethods to determine the concentration of electrolytes and blood acid-base status promoted physiological and clinical research into acid-base metabolism in the 1960s. In the new physiologically based terminology of systemic acid-base status, the relationship between blood acid-base status and net acid intake or excretion was, however, incorrectly simplified. In the 1970s metabolic acidosis was observed in patients on chemically defined diets and parenteral nutrition. Based on the data of comprehensive acid-base balance studies, calculation models were used to estimate renal net acid excretion from nutrient intake and to predict the potential renal acid load of single foods. Extrapolating current trends to the future, one can say that acid-base physiology will probably remain a challenge in nutrition and functional medicine over the next few years. The challenge will include new concepts for the manipulation of nutritional acid load in sports, dietetics and preventive medicine as well as new definitions of the upper intake level of potential renal acid load in functional foods and the monitoring of renal net acid excretion in populations. Received: 12 March 2001, Accepted: 23 March 2001     

Nutrition, acid-base status and growth in early childhood

Summary Optimal growth is only possible in a well-balanced “inner milieu”. Premature infants are especially vulnerable for disturbances of acid-base metabolism with a predisposition to metabolic acidosis due to a transient disproportion between age-related low renal capacity for net acid excretion (NAE) and an unphysiologically high actual renal NAE on nutrition with standard formulas. During a 50 month period, 452 low birth-weight infants were screened for spontaneous development of incipient late metabolic acidosis (ILMA), an early stage during the development of retention acidosis, characterized by maximum renal acid stimulation (MRAS, urine-pH < 5.4) on two consecutive days but still compensated systemic acid-base status. Compared with controls, patients with ILMA showed higher serum creatinine values, an increased urinary excretion of sodium, aldosterone and nitrogen, but only slightly lower blood pH (7.38 vs 7.41) and base excess (−2.8 vs. 0.2 mmol/l) with respiratory compensation (PCO₂ 35 vs 37 mm Hg). Patients with altogether 149 episodes of ILMA were subsequently randomly allocated to either treatment with NaHCO₃ 2 mmol/kg/d for 7 days or no special therapy in protocol I, or NaHCO₃ vs NaCl each 2 mmol/kg/d for 7 days in protocol II. Patients of protocol I with persistent MRAS for 7 days showed lowest weight gain and a tendency for a further increase in urinary aldosterone and nitrogen excretion. NaCl supplementation (protocol II) seemed to promote weight gain without affecting either impaired mineralization or suboptimal nitrogen retention. Patients with alkali therapy under both protocols showed normal weight gain and normalization of hormonal stimulation, mineralization (protocol II) and nitrogen assimilation. Modification of the mineral content of a standard preterm formula decreased renal NAE to the low level seen on alimentation with human milk and reduced the incidence of ILMA in preterm and small-for-gestational-age infants to 1 %. The data show that ILMA is associated with impaired growth. Activation of secondary homeostatic mechanisms (extracellular volume contraction, depletion of disposable net base pools) might be important for impaired growth. Production of new formulas for reduced renal NAE could be an effective general preventive measure to reduce the clinical importance of one component of mixed acid-base disorders in early childhood. Received: 26 February 2001, Accepted: 7 March 2001     

The acid-base hypothesis: diet and bone in the Framingham Osteoporosis Study

Summary Background There continues to be considerable debate about the role of acid vs. basic components of the diet on the long-term status of bone mineral density. Aim In a set of two analyses, we examined the effect of components in the diet thought to have basic effects (magnesium, potassium, fruit, vegetables) and acid effects (protein) on bone mineral density in an elderly cohort. Methods Bone mineral density of participants in the Framingham Osteoporosis Study was measured at three hip sites and one forearm site at two points in time, four years apart. At the time of baseline measurement, participants ranged in age from 69–97 years. Dietary intake was assessed at baseline by food frequency questionnaire. Results As hypothesized, magnesium, potassium, fruit and vegetable intakes were significantly associated with bone mineral density at baseline and among men, with lower bone loss over four years. In contrast to the hypothesis, higher rather than lower protein intakes were associated with lower bone loss. Conclusion Together these results support the role of base forming foods and nutrients in bone maintenance. The role of protein appears to be complex and is probably dependent on the presence of other nutrients available in a mixed diet. A balanced diet with ample fruit and vegetables and adequate protein appears to be important to bone mineral density. Received: 4 August 2001, Accepted: 14 August 2001     

Plasma levels of antioxidant vitamins and cholesterol in a large population sample in Central-Northern Italy

Background: In a population-based multicenter case-control study of diet, life-style, and gastric cancer a large series of adults, aged 30–75 years (mean 58.9 yearss), were randomly sampled from the general population in 3 areas of Central-Northern Italy. Aim of the study: To evaluate the relationship between plasma levels of antioxidant vitamins and cholesterol, and sociodemographic characteristics, life-style factors, and dietary intake of selected nutrients in a sample of the Italian population. Methods: A fasting blood sample was available for 945 subjects (553 men, 392 women). The plasma concentrations of ascorbic acid, carotene, retinol, alpha-tocopherol, and cholesterol were determined by a centralized laboratory. All participants answered to a detailed questionnaire collecting information on sociodemographic and anthropometric characteristics, smoking, alcohol drinking, and dietary habits. Covariance analysis models, with post hoc Dunnett tests, including terms for age, sex, study center, and period of blood drawing, were used for selected multiple-way comparisons of mean values of plasma nutrients. Results: Mean plasma values of retinol were higher among men while women had higher levels of plasma carotene, ascorbic acid, alpha-tocopherol, and cholesterol. Plasma carotene levels showed an inverse association with body mass index, alcohol consumption, and smoking and a positive association with social class. Carotene concentrations were higher in plasma samples obtained in spring/summer, while ascorbic acid levels were higher in autumn/winter. Partial correlation coefficients between carotene and ascorbic acid (0.69 in men; 0.74 in women), between carotene and alpha-tocopherol (0.44; 0.37), and between alpha-tocopherol and ascorbic acid (0.45; 0.41). Plasma alpha-tocopherol and retinol correlated with plasma cholesterol. On the other hand, plasma carotene and ascorbic acid were correlated with their estimated dietary intakes, while the intakes of other nutrients, as expected, correlated rather poorly with the respective plasma concentrations. Conclusions: Socio-economic factors, life-style, and specific nutrient intake, in addition to gender, are related to nutrient plasma levels in Italian adults and may provide specific suggestions for the prevention of chronic diseases. Received: 8 December 1998, Accepted: 25 February 1999     

Dietary lean red meat and human evolution

Summary Scientific evidence is accumulating that meat itself is not a risk factor for Western lifestyle diseases such as cardiovascular disease, but rather the risk stems from the excessive fat and particularly saturated fat associated with the meat of modern domesticated animals. In our own studies, we have shown evidence that diets high in lean red meat can actually lower plasma cholesterol, contribute significantly to tissue omega-3 fatty acid and provide a good source of iron, zinc and vitamin B₁₂. A study of human and pre-human diet history shows that for a period of at least 2 million years the human ancestral line had been consuming increasing quantities of meat. During that time, evolutionary selection was in action, adapting our genetic make up and hence our physiological features to a diet high in lean meat. This meat was wild game meat, low in total and saturated fat and relatively rich in polyunsaturated fatty acids (PUFA). The evidence presented in this review looks at various lines of study which indicate the reliance on meat intake as a major energy source by pre-agricultural humans. The distinct fields briefly reviewed include: fossil isotope studies, human gut morphology, human encephalisation and energy requirements, optimal foraging theory, insulin resistance and studies on hunter-gatherer societies. In conclusion, lean meat is a healthy and beneficial component of any well-balanced diet as long as it is fat trimmed and consumed as part of a varied diet. Received: 24 February 2000, Accepted: 20 March 2000     

Minimal-invasive approach to study pulmonary, metabolic and renal responses to alimentary acid-base changes in conscious rabbits

Summary Background Systemic acid-base balance is maintained by the complex interplay of renal and pulmonary control functions and metabolic adaptations, whereby intake and mineral composition of feed are important factors. Aim of the study It was intended to explore the role of alimentary acid-base load and carbonic anhydrase activity for regulatory responses of renal, pulmonary or metabolic origin in rabbits as typical herbivores. Methods Sixty-eight conscious male rabbits (about 3.5 kg) were kept in a metabolic cage, to determine daily water intake, urine excretion and food consumption. Different groups were fed either alkali-rich rabbit standard pellets, or modified rabbit chow with low Ca⁺⁺-content, or a special diet with very low alkali content, or standard food together with a low oral dose (about 20 mg · kg⁻¹· d⁻¹) acetazolamide. Samples from the central ear artery were analyzed for blood gases (PaO₂, PaCO₂), pHa, base excess (BE) and actual bicarbonate (HCO⁻ _3a). The metabolic CO₂ production (CO_{2 STPD}) was determined, to calculate alveolar ventilation ( _{A BTPS}). Anaerobically collected urine was analyzed for pHu and for concentrations of bicarbonate/carbonate (HCO⁻ ₃/CO⁻⁻ ₃), ammonium (NH₄ ⁺), and phosphate. Results 1) Systemic BE was not affected by alimentary alkali load, either varied spontaneously by standard food intake or by the low-Ca⁺⁺ diet, and decreased only slightly on the low-alkali diet, but distinctly upon carbonic anhydrase inhibition. 2) Under all conditions of alimentation, PaCO₂ was closely correlated with BE without a detectable set-point, the normal-range variability of BE being sufficient to elicit corresponding changes in _A. In contrast, acetazolamide led to much lower values of PaCO₂ than predicted by the reference PCO₂/BE relationship, being primarily caused by significant reductions in CO₂ (> 20 %). 3) Prior to other systems, renal base excretion, normally being high on species-adapted standard chow, closely followed any variation of alimentary alkali load and approached zero upon the low-alkali diet. It was, however, not significantly influenced by carbonic anhydrase (CA) inhibition on alkali-rich alimentation. Conclusions Blood acid-base balance in rabbits is maintained over a wide range of alimentary alkali load by effective adaptation of renal base excretion, independent of CA activity. Ventilatory pH control is perpetuated even in the normal range of BE, provided metabolic rate is not impaired, e. g., by CA inhibition. These results may help one understand the different manifestations of acid-base disorders in body fluids under clinical conditions. Received: 7 March 2001, Accepted: 15 March 2001     

Animal nutrition and acid-base balance

Summary In domestic animals, acid-base balance may be influenced by nutrition. The major research effort in this area has been made on the prevention of hypocalcemic postparturient paresis in dairy cows. This disorder is caused by the sudden increase of calcium secretion into the colostrum. The manipulation of the dietary cation-anion difference makes it possible to maintain the cows in metabolic acidosis during the critical period that precedes calving, presumably via a mechanism that involves the strong ion difference in the extracellular fluid. As a consequence the mobilization of calcium is enhanced and the incidence of the disorder is decreased. Conversely, a dietary induced metabolic alkalosis leads to a more severe degree of hypocalcemia and the incidence of the disease is increased. The underlying mechanisms of the prevention are only partially understood. Nevertheless, this preventive method is already widely applied in practice. Nutrition effects on acid-base balance also influence growth and food intake in higher vertebrates and fish. As a consequence, the incidence of developmental orthopedic diseases in fast-growing domestic animal species may be affected. Also, the bone mineral content of athletic horses may be influenced by dietary induced modification of the acid-base status. The mineral loss due to metabolic acidosis may lead to an increase in the incidence of stress fractures. This overview should give insight into relevant aspects of nutrition and acid-base balance in domestic animal species. Received: 28 March 2001, Accepted: 10 April 2001     

Diet restriction in mice causes differential tissue responses in total reducing power and antioxidant compounds

Summary Background: Diet restriction (DR) has been shown to extend the life spans of various laboratory animals, the mechanism may involve a decrease in oxidative stress. When determining if the total tissue defense has been altered, it is important to observe the overall direct antioxidant capacity, which consists of low molecular weight antioxidants (LMWA) and enzymes. Aim: To determine DR induced changes in total reducing power and overall direct antioxidant capacity of various mouse tissues. Methods: Young female Sabra mice were fed a 60% food restricted diet for 40 days (DR group). Organs of the DR group and of ad libitum (AL) fed controls were then dissected and examined. A cyclic voltammetry method was used to quantify the total reducing power, which correlates with the overall LMWA activity. Specific LMWA were identified by HPLC-ECD. Superoxide dismutase activity and H₂O₂ degrading ability were measured in order to include the enzymatic antioxidant component. Results: Short-term DR caused alterations in the total reducing power of various mouse tissues, indicating changes in the total scavenging ability of these tissues. Overall direct antioxidant capacity of heart, kidney and muscle was enhanced; liver and small intestine deteriorated; brain did not differ between DR and AL groups; lung and spleen exhibited a mixed response. Conclusions: We have shown for the first time that DR causes changes in the total reducing power of different mouse tissues, thus, affecting the overall direct antioxidant capacity. These findings support the suggestion that there may be a biological regulation of the antioxidant system. Received: 31 March 1999, Accepted: 11 February 2000     

Effects of maternal chronic alcohol administration in the rat: lactation performance and pup's growth

Summary A fostering/crossfostering analysis of the effects of maternal ethanol exposure on lactation performance and offspring growth was performed. Wistar rats were kept under one of the three experimental nutritional treatments: alcohol-treated (EG), pair-fed-treated (PFG) (as a nutritional control of alcohol-associated malnutrition), and control or normal diet (CG). Rats from the EG group were accustomed to increased amounts of ethanol (5 % during the first week to 20 % in the fourth week). The 20 % ethanol level was maintained throughout three additional weeks and during gestational and lactational period. Daily food intake, fluid consumption, body weight and gestational parameters were studied in control (CG), pair-fed (PFG) and ethanol dams (EG). At birth, half the litters were fostered to other dams of the same treatment (GLG) and half were cross-fostered to dams of the opposite treatment (GG, LG). No cross-fostering analyses were performed on the pair-fed group. Offspring body weight was controlled throughout lactation. Liver, kidney and spleen weights as well as milk consumption were also studied at the end of lactation period. In dams, a significant reduction of body weight was described throughout the suckling period. No ethanol detrimental effects were observed on body weight at birth, but in spite of a normal birth weight, alcohol during lactation was responsible for a growth deficit. Milk consumption was significantly reduced in offspring exposed to ethanol during gestation and/or lactation. Curiously, prenatal alcohol exposure affects adversely the suckling behaviour in pups at the time of weaning. In our study, alcohol treatment and malnutrition affects liver and spleen weights. However, malnutrition decreases spleen weights more than alcohol treatment. In the case of the kidney weights the alcohol decreases kidney weight more than malnutrition. Collectively, the data from the present study show similar effects following pre/postnatal and postnatal alcohol exposure. The findings suggest that chronic alcohol administration during gestation and/or lactation adversely affects pup growth at weaning as indicated by its effect on milk consumption, pup and organ weight. Received: 3 January 2001, Accepted: 8 June 2001     

Effect of lactobacilli, bifidobacteria and inulin on the formation of aberrant crypt foci in rats

Summary Background Our studies were aimed at investigating the effect of lactic acid producing bacteria (LAB) or inulin, a natural source of non-digestible oligosaccharides derived from chicory, on the induction by carcinogens of aberrant crypt foci (ACF) in the colon, which are considered to be early precursor lesions of neoplasia. Methods Strains of Bifidobacterium longum, Lactobacillus casei and Lactobacillus acidophilus were administered to rats fed a purified high starch diet, under a variety of treatment protocols including daily gavage, via the drinking water and in the diet. The rats were treated with methyl-N-nitrosourea, 1,2-dimethylhydrazine, or azoxymethane (AOM) to induce ACF. Results In general, no consistent significant changes in ACF numbers were detected in these experiments. In one study, the basal diet of the rats was changed to one containing a higher level of fat (corn oil). Under these conditions, a significant decrease in AOM-induced colonic ACF was seen in rats given L. acidophilus or inulin. In a concurrent group of animals fed a low fat diet, no significant decrease in ACF was observed. Conclusions The results indicate that the type of diet fed can influence the detection of protective effects of LAB and oligosaccharides and that against the background of a diet with a level of fat typical of a Western diet, evidence for a protective effect of L. acidophilus and inulin towards colon cancer was obtained Received: 26 July 2001, Accepted: 26 November 2001     

Small supplements of N-3 fatty acids change serum low density lipoprotein composition by decreasing phospholipid and apolipoprotein B concentrations in young adult women

Summary In order to investigate the effect of a short-term application of marine n-3 polyunsaturated fatty acids on the composition of serum very low density lipoproteins (VLDL), low density lipoproteins (LDL), and high density lipoproteins (HDL), nine women aged 29±4.2 years, following a diet with a SFA/MUFA/PUFA profile of 2.4/3/1, received supplements of six capsules daily, each capsule containing 0.137 g of n-3 fatty acids (14.5% eicosapentaenoic acid (EPA) and 8.9% docosahexaenoic acid (DHA)) for 10 d. Food consumption, assessed during two 10-days periods indicates that percentage contribution of SFA, MUFA, and PUFA to the daily energy intake did not change through the fish-oil supplementation period, but the daily consumption of n-3 fatty acids increased 2.3 times. N-3 fatty supplementation increased EPA and DHA percentages in serum phospholipids, but failed to decrease (p>0.05) the cholesterol and triglyceride concentration in serum LDL and HDL, although it did so in VLDL. In contrast, the lipoprotein-phospholipid and lipoprotein-protein concentrations were markedly affected, mainly in LDL and HDL (at least p<0.01). HDL and VLDL compositions were not affected but the total mass (lipid+protein in mg/dl) concentration of these lipoproteins significantly decreased (p<0.05), suggesting a lower number of these particles in circulating blood after the n-3 treatment. The LDL-cholesterol/LDL-apolipoprotein B ratio increased (p<0.01) reflecting a probable increase in LDL size. Following fish oil supplementation, LDL particles contained a significantly lower amount of phospholipids, which also suggests changes in the surface/core ratio of the average LDL. Changes in serum lipoprotein lipids did not significantly correlate with any dietary change other than the n-3 fatty acid increase. The results indicate that a 10-day application of a small supplement of n-3 change the LDL composition leading to less atherogenic LDL particles with lower phospholipid and apolipoprotein (Apo) B concentrations. Received: 15 May 1998, Accepted: 28 August 1998     

Hyperhomocysteinemia,and low intakes of folic acid and vitamin B12 in urban North India

Summary Background and Aim An adverse coronary risk profile has been reported amongst rural-to-urban migrant population living in urban slums undergoing stressful socio-economic transition. These individuals are likely to have low intakes of folic acid and vitamin B12, which may have an adverse impact on serum levels of homocysteine (Hcy). To test this hypothesis, we studied serum levels of Hcy in subjects living in an urban slum of North India and healthy subjects from urban non-slum area. Methods Group I consisted of 46 subjects (22 males and 24 females) living in an urban slum, while group II consisted of healthy subjects (n = 26, 13 males and 13 females) living in the adjacent non-slum area. Anthropometric measurements, biochemical profile (fasting blood glucose, total cholesterol, serum triglycerides, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol) and fasting serum levels of Hcy were measured. Dietary intakes of folic acid, vitamin B12, vitamin B1, and iron were calculated by the 24-hour dietary recall method. Serum levels of Hcy were correlated with dietary intakes of nutrients, anthropometry, and metabolic variables. Results Sex-adjusted serum levels of Hcy in mmol/L (Mean ± SD) were high, though statistically comparable, in both the groups (group I: 20.8 ± 5.9 and group II: 23.2 ± 5.9). Overall, higher than normal serum levels of Hcy (> 15 μmol/L) were recorded in 84 % of the subjects. A substantial proportion of subjects in both groups had daily nutrient intakes below that recommended for the Asian Indian population (folic acid: 93.4 % in group I and 96.7 % in group II, vitamin B12: 76.1 % in group I and 88.4 % in group II). However, between the two groups, average daily dietary intakes of both the nutrients were statistically comparable. As compared to non-vegetarians, vegetarians showed lower intakes of folic acid (p < 0.01) and vitamin B12 (p < 0.01) in both groups. On multivariate linear regression analysis with serum Hcy as the response variable and vegetarian/non-vegetarian status and sex (male/female) as predictor variables, higher serum levels of Hcy were observed in vegetarians vs non-vegetarians (β = 4.6, p < 0.05) and males vs females (β = 5.3, p < 0.01). Conclusions Low intakes of folic acid and vitamin B12, and hyperhomocysteinemia, in both the healthy population living in urban slums and adjacent urban non-slum areas, are important observations for the prevention of nutritional and cardiovascular diseases in the Indian subcontinent. Received: 30 October 2001, Accepted: 14 January 2002     

Nutrient oxidation and metabolic rate as affected by meals containing different proportions of carbohydrate and fat, in healthy young women

Summary The amount and composition of food eaten influence body weight regulation, which requires that, in the long term, energy intake matches energy expenditure and that the oxidation rate to be equal to intake for each and separate nutrients. The aim of this study was to examine the influence of two liquid formulas with different macronutrient composition, a high carbohydrate (HC) meal as compared to a high fat (HF) meal, on substrate oxidation and on thermic effect of food (TEF). Eighteen lean and healthy women which were fed a HC diet during the 3 preceding d were studied for a further 4 h after meal intake. The test meals provided fixed energy intake and whose calculated FQ were 0.77 for HF meal, and 0.96 for HC meal. The mean NPRQs were higher (P<0.01) in the HC group than in the HF group, even with values greater than 1.00 indicating net lipid synthesis (NL), and which correlated with metabolic rate (MR) value (P<0.05), glucose (P<0.05), and heart rate (HR) values. Carbohydrate (CHO) oxidation was higher with the HC than with HF meal (P<0.01) and correlated with the MR (P<0.05). Protein oxidation rate rose above baseline (P<0.01); this increase was accompanied by with a negative CHO balance. It is concluded that the change in fuel selection and the increase of TEF is mainly due to CHO intake and metabolism, respectively, and that surplus of dietary CHO of preceding days together with a large load of CHO can exceed the glycogen storage capacity and trigger NL. Received: 29 September 1998, Accepted: 14 April 1999     

Green tea extract decreases plasma malondialdehyde concentration but does not affect other indicators of oxidative stress, nitric oxide production, or hemostatic factors during a high-linoleic acid diet in healthy females

Summary Background: Green tea contains polyphenolic catechins which can act as antioxidants and thus decrease the risk for cardiovascular diseases. Aim of the study: To investigate whether green tea extract differs from placebo in its effects on markers of antioxidant status, lipid peroxidation, nitric oxide production, thromboxane production, and blood coagulation during a controlled high linoleic acid diet in healthy subjects. Methods: Twenty healthy non-smoking females (23–50 years) participated in a 4-week controlled intervention study. The experimental diet was rich in linoleic acid (9 en%) and contained fat, protein and carbohydrates: 27, 14, and 59 en%, respectively. In addition, the subjects ingested encapsulated green tea extract (3 g/d) or placebo mixture in a double-blind manner. Fasting blood samples and five 24-hour urines were collected before and at the end of the 4-week experimental period. Same samples were received from 10 control subjects. Results: Green tea extract significantly decreased plasma malondialdehyde (MDA) concentration in comparison with the placebo treatment. The treatments did not differ in serum lipids, indicators of antioxidant status, urinary 8-iso-prostaglandin F_2α, 2,3-dinor-thromboxane B₂, nitric oxide metabolites or coagulation indicators. Conclusions: We conclude that an amount of green tea extract which corresponds to 10 cups of tea per day for 4 weeks does not have specific effects on several indicators related to risk of cardiovascular diseases in comparison with placebo treatment. The relatively small but significant decrease in lipid peroxidation indicated by decreased plasma MDA was not associated with changes in markers of oxidative stress (urinary 8-iso-prostaglandin F_2α and blood oxidized glutathione) or hemostasis. Received: 8 January 1999, Accepted: 12 April 1999     

Short-term fish oil supplementation improved innate immunity, but increased ex vivo oxidation of LDL in man--a pilot study

Summary Background: Fish and fish oils are rich in the two long-chain polyunsaturated fatty acids (LCPUFAs) eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3). The n-3 LCPUFAs have been reported to have beneficial effects on cardiovascular functions, but their role in relation to immune functions is still controversial. Aim of the study: The objectives of this study were to determine the effects of supplementation with fish oil on immune cell functions in human subjects. We have also assessed the effects on plasma lipids, antioxidant status and susceptibility of low-density lipoproteins (LDL) to oxidative stress. The antioxidant status was determined by measuring plasma vitamin C, tocopherols and carotenoids in plasma and LDL, and superoxide dismutase (SOD) in red blood cells. Design: For 30 days, 10 volunteers ingested 25 g/d of either fish oil, providing n-3 LCPUFAs (7.5 g), or high-oleic sunflower oil, providing monounsaturated fatty acids mainly as oleic acid (22 g). The oils contained similar profiles of tocopherols. At day 0 and day 30, blood samples were drawn by venipuncture for plasma lipid and antioxidant analyses and lipoprotein isolation, and for isolation and functional tests of mononuclear cells and granulocytes. Fatty acid profiles of immune cells and LDL were also determined. Results: Fish oil supplementation resulted in an accumulation of n-3 LCPUFAs (EPA, DHA) in LDL and immune cells. The phagocytic activity, a measure of immune cell activity, was increased in both groups. Whereas the plasma and LDL antioxidant status do not appear to be affected by fish oil supplementation, an increased susceptibility of LDL to oxidation was observed in these healthy volunteers. Conclusions: The optimal amounts of n-3 fatty acids required to modulate immune functions remain to be established. In addition, adequate levels of antioxidant protection need to be provided during fish oil supplementation. Received: 19 September 2000 / Accepted: 12 February 2001     

Vegetables, fruit, antioxidants and cancer: a review of Italian studies

Summary Background Case-control studies have suggested that a diet rich in fresh fruit and vegetables protects from the risk of most common epithelial cancers, including those of the digestive tract, and also several nondigestive neoplasms; however, selections in cohort studies have been generally weaker. Aim of the study To review the relation between frequency of consumption of vegetables and fruit, estimated intake of selected antioxidants and the risk of cancer at different sites. Methods Systematic overview of data, with specific focus on a network of case-control studies conducted in Italy from 1983 to 1999. Results The relative risks (RR) of digestive tract neoplasms were reduced in subjects reporting highest vegetable intake. A protective effect of vegetables was also observed for hormone-related neoplasms. Fruit was related to a reduced RR of cancers of the upper digestive tract, stomach and urinary tract. With reference to the role of selected antioxidants, beta-carotene, vitamins C and E showed a significant inverse relation with oral and pharyngeal, esophageal and breast cancer risk. Against colorectal cancer, the most consistent protective effects were provided by carotene, riboflavin and vitamin C, but inverse relations were observed also for calcium and vitamin D. Conclusions Fruit and vegetable consumption in Mediterranean populations appears to provide protection against several types of neoplasms. Received: 27 July 2001, Accepted: 23 October 2001     

北京市小学生与肥胖相关知识、态度、行为的现况调查

目的　了解北京市小学生的营养状况及其膳食与运动相关知识、态度、行为的现状。方法　采用整群抽样法抽取北京市 4所小学三、四年级全体学生 ,进行问卷调查。结果　北京市小学生瘦长型营养不良率1 3 0 3 % ,肥胖型营养不良率 3 2 0 4%。膳食与运动相关知识掌握较好的学生占 0 73 % ,正确态度持有率 5 6 75 % ,良好行为形成率 1 1 2 7%。知信行均为女生好于男生。知信行间存在一定的关联。结论　健康教育可传授正确的膳食与运动知识给学生 ,转变他们的错误态度 ,从而改变他们的不良行为习惯 ,以促进学生营养状况的改善。     

Effects of insulin and amino acids on leucine metabolism in young and middle-aged humans

Summary Background Aging is characterized by loss of muscle mass. In healthy subjects this process is associated with hormone and nutritional changes which take place over many decades. Aim of the study To investigate the effects of insulin and amino acids on amino acid metabolism in middle-aged humans. Methods We evaluated leucine kinetics by means of the intravenous infusion of [1-¹⁴C]leucine, in 8 young (age 24±2 yr, BMI 21±2 kg/m²) and in 6 middle-aged (age 53±4 yr, BMI 26±1 kg/m²) healthy subjects. Studies were performed under fasting conditions (basal), and after 180 min of euglycemic hyperinsulinemic clamp (study I), or 180 min of euglycemic hyperinsulinemia in combination with an intravenous amino acid infusion (study II). Results In the basal state endogenous leucine flux (ELF, an index of proteolysis), normalized for IBW, averaged 1.71±0.12 and 1.66±0.14 μmol/kg·min in young and middle-aged subjects, respectively. Basal leucine oxidation (0.22±0.03 vs 0.28±0.03 μmol/kg·min, p < 0.05) was lower in middle-aged with respect to young subjects. Non-oxidative leucine disposal (NOLD, an index of protein synthesis: 1.44±0.11 vs 1.43±0.11 μmol/kg·min) was similar in young and middle-aged subjects, respectively. In response to insulin (study I) the absolute and percent decline of ELF and LOX were similar in young and middle-aged subjects: ELF declined to 1.05±0.06 μmol/kg·<,min (−M39±5 %) and 1.07±0.14 μmol/kg·min (−36±4 %), in young and middle-aged, respectively (both p < 0.01 vs basal); LOX declined to 0.21±0.02 μmol/kg·min (−35±3 %), and 0.18±0.05 μmol/kg·min (−28±3 %, p < 0.05 vs basal) in young and middle-aged individuals respectively (both p < 0.01 vs basal). In contrast, insulin-mediated whole-body glucose uptake was lower in middle-aged subjects (6.6±1.4 mg/kg·min) with respect to young individuals (8.1±1.7 mg/kg·min, p < 0.05). During study II (insulin plus AA) a significant rise in NOLD was obtained in both young (1.72±0.10 μmol/kg·min, p < 0.01 vs basal) and middle-aged subjects (1.76±0.25 μmol/kg·min, p < 0.01 vs basal). Similarly, net leucine balance rose significantly in both young (+0.62±0.13 vs −0.25±0.02 μmol/kg·min, p < 0.01 vs basal) and middle-aged subjects (+0.37±0.08 vs −0.22±0.03 μmol/kg·min, p < 0.01 vs basal) suggesting that the anabolic response to amino acids is preserved in middle-aged subjects. Conclusions In middle-aged subjects we observed 1) a moderate decline in basal leucine oxidation; 2) a normal antiproteolytic response to insulin and a reduction in glucose uptake; and 3) a normal anabolic response to AA plus insulin. In conclusion, the data provide evidence for a normal regulation of protein anabolism and an early dissociation between the metabolic effects of insulin on glucose uptake and proteolysis in middle-aged subjects. Received: 6 September 2000, Accepted: 10 July 2001     

Pollen-related food allergy: cloning and immunological analysis of isoforms and mutants of Mal d 1, the major apple allergen, and Bet v 1, the major birch pollen allergen

Summary Background: Mal d 1, the major apple allergen, cross-reacts with IgE specific for the major birch pollen allergen, Bet v 1, and is responsible for birch pollen related food allergy to apple. Isoforms of Bet v 1 showing minor sequence variations display different binding capacitiy for specific IgE antibodies from allergic patients. Moreover, strain-dependent variation of allergenicity has been reported for apples. Objective: To investigate the occurence of strain-dependent isoforms of Mal d 1 which may differ in their allergenic potential, to obtain data on structures essential for binding of Mal d 1 to the antibody, and to gain insights into the structures responsible for its IgE cross-reactivity to Bet v 1. Methods: The cDNA of Mal d 1 from various apple strains was amplified by a PCR strategy based on conserved regions of known Mal d 1-sequences, and sequenced. Two major isoforms of Mal d 1were expressed as recombinant proteins and purified, as were different variants of the major birch pollen allergen, Bet v 1. Together with already existing recombinant birch pollen and apple allergens, these were subjected to allergenicity testing by IgE-immunoblotting, enzyme allergo sorbent test and dose related mediator release. “Hot-spots” for IgE-reactivity were identified by site-directed mutagenesis. Results: Twelve Mal d 1-clones were sequenced from 7 apple varieties and compared to 3 known Mal d 1 sequences. The clones were clustered into two groups, each showing a high degree of sequence identity to one of the known sequences and specific differences to the third sequence. No strain-specific sequences were identified. In contrast, apple strains with reported differences in allergenicity showed different expression levels of the major allergen. Immunologic testing of recombinant allergens revealed high IgE binding capacity of 2 major isoforms, named GD26 and GS29, with a slightly higher IgE binding capacity of DG26. Moreover, the allergenicity was similar to another rMal d 1 reported in the literature, representing the isoform divergent from our clones. Mutational analysis of our Mal d 1 allergens identified serine in position 111 as essential for IgE binding. Allergenicity was almost depleted by changing this residue into a proline. Moreover, the corresponding serine residue, present in position 112 of Bet v 1, was in a similar manner crucial for the allergenicity of the birch pollen allergen. Conclusion: We conclude that divergent allergenicity of apple strains mainly depends on differnet expression levels of the major allergen. Introduction of a proline residue in position 111 of Mal d 1 and in position 112 of Bet v 1 led to a drastic reduction of allergenicity of both the pollen and the food allergen, obviously also removing the cross-reactive epitope. Mutants with reduced IgE-reactivity but maintained T-cell reactivity may represent new candidates for a safer specific immunotherapy with reduced side-effects. Received: 7 June 1999, Accepted: 30 July 1999