Knowledge of the cannabinoid system and its components has expanded greatly over the past decade. There is increasing evidence for its role in the regulation of food intake and appetite. Cannabinoid system activity in the hypothalamus is thought to contribute to the homeostatic regulation of energy balance, under the control of the hormone leptin. A second component of cannabinoid-mediated food intake appears to involve reward pathways and the hedonic aspect of eating. With the cannabinoid system contributing to both regulatory pathways, it presents an attractive therapeutic target for the treatment of both obesity and eating disorders.

The lipase inhibitor, orlistat, is used in the treatment of obesity and reduces fat absorption by about 30%. However, the mean weight loss induced by orlistat is less than expected for the degree of fat malabsorption. It was hypothesised that lipase inhibition with orlistat attenuates the suppressive effects of oral fat on subsequent energy intake in normal-weight subjects. Fourteen healthy, lean subjects (nine males, five females; aged 25±1·3 years) were studied twice, in a double-blind fashion. The subjects received a high-fat yoghurt ‘preload’ (males 400g (2562kJ); females 300g (1923kJ)), containing orlistat (120mg) on one study day (and no orlistat on the other ‘control’ day), 30min before ad libitum access to food and drinks; energy intake was assessed during the following 8h. Blood samples were taken at regular intervals for the measurement of plasma cholecystokinin (CCK). Each subject performed a 3d faecal fat collection following each study. Energy intake during the day was greater following orlistat (10220 (sem 928) kJ) v. control (9405 (sem 824) kJ) (P=0·02). On both days plasma CCK increased (P<0·05) after the preload. Plasma CCK 20min following ingestion of the preload was less after orlistat (4·1 (sem 0·9) pmol/l) v. control (5·3) (sem 0·9) pmol/l (P=0·028); however there was no difference in the area under the curve 0–510min between the two study days. Fat excretion was greater following orlistat (1017 (sem 168) kJ) v. control (484 (sem 90) kJ) (P=0·004). In conclusion, in healthy, lean subjects the acute inhibitory effect of fat on subsequent energy intake is attenuated by orlistat and the increase in energy intake approximates the energy lost due to fat malabsorption.

The discovery of the production of leptin by the stomach, in addition to its production by adipose tissue, has initiated new investigation into the possible role of this protein in the digestive physiology, in particular in the short-term control of energy balance. Leptin has been identified in the lower half of the stomach glands both in the pepsinogen granules of chief cells and in the granules of a specific endocrine cell type, suggesting that leptin action is exerted by both exocrine and endocrine pathways. Gastric leptin is sensitive to the nutritional state, being rapidly mobilized in response to food intake following fasting, or after the administration of satiety factors; this suggests a role for this protein in the short-term regulation of feeding, acting in collaboration with satiety peptides such as cholecystokinin. Leptin, produced by gastric cells and by adipocytes, could act on both acute and chronic regulation of feeding behaviour respectively, giving information to the brain on the availability of external (food) and internal (fat depots) energy resources, thus participating in short- and long-term satiation.

The objective of the present study was to investigate the effects of starches with differing rates of hydrolysis on exposure to pancreatin in vitro on postprandial carbohydrate metabolism in healthy subjects and in subjects with type 2 diabetes. Two test starches, prepared from uncooked native granular starch products, and naturally enriched with 13C, were consumed in a randomized crossover design by eight healthy and thirteen type 2 diabetic subjects. One starch was characterized in vitro as being rapidly hydrolysed (R, 94% after 180min), and the other was more slowly hydrolysed (S, 51% after 180min). Each subject consumed 50g of each test starch. In addition, the type 2 diabetic subjects consumed 89·7g of the S starch on a separate occasion. Blood samples were taken at 10min intervals for 3h, and at 20min intervals for a further 3h during a 6h postprandial period. Breath 13CO2 enrichment was measured at the same time points, and indirect calorimetry was performed for seven 20min sessions immediately before and during the 6h postprandial period. With the R starch, plasma glucose concentrations and serum insulin concentrations rose faster and the maximum glucose change was approximately 1·8 times that for the S starch, averaged across both subject groups. The areas under the curves for glucose and insulin were, respectively, 1·7 and 1·8 times higher for the R starch compared with the S starch, averaged across both subject groups. The rate of 13CO2 output and the proportion of 13C recovered in breath after consumption of the R starch was similar for both subject groups. The results provide evidence that starches which have different rates of hydrolysis in vitro result in different patterns of glycaemia and insulinaemia in both healthy adults and in diet-controlled type 2 diabetic subjects. Data from the hydrolysis of novel starch products in vitro, therefore, are useful in predicting glycaemic responses in vivo.

Lack of breakfast has been implicated as a factor contributing to children's poor diets and school performance. Breakfast-club schemes, where children are provided with breakfast in school at the start of the school day, have been initiated by the Department of Health in schools throughout England, UK. The aim of the present study was to compare the energy and nutrient intakes of schoolchildren who attended breakfast clubs (attendee subjects) with those who did not (control subjects). Three different schools were studied, involving a total of 111 children aged between 9 and 15 years. There were fifty-nine attendee and fifty-two control subjects. The two groups were matched for eligibility for school meals. All subjects completed a 3 d weighed food diary for estimation of nutrient intake. Height and weight were measured and BMI calculated. Nutrient intake data were analysed using a general linear model with age as a covariate. The demographic and anthropometric characteristics of the attendee and control subjects were similar. Children who attended breakfast clubs had significantly greater intakes of fat (% energy), saturated fat (% energy) and Na than control subjects. Thus, in these schools breakfast-club participation was not associated with superior nutrient intake or improvements in dietary pattern.

We studied the effects of α-tocopheryl acetate supplementation on the development of fatty streaks and its ability to modulate the expression of monocyte chemoattractant protein (MCP)-1 in aortic lesions of apoliprotein E knockout mice. For this purpose, 16-week-old apolipoprotein E knockout mice received α-tocopherol supplemention ((800 mg)/kg diet) for 6 weeks. After this time, total and lipoprotein cholesterol in the serum, hepatic tocopherol, aortic lesion area and MCP-1 (protein and mRNA) expression were analysed. Our present results showed that the dietary supplementation with α-tocopherol did not reduce serum cholesterol nor change lipoprotein profile, but it reduced the area of the aortic lesion by 55%. The reduction in the lesion size was correlated with the reduced expression of MCP-1 mRNA and protein, as detected by real-time quantitative polymerase chain reaction and immunohistochemistry respectively. In conclusion, the results obtained here are relevant to the study of atherosclerosis, as they correlate the effectiveness of vitamin E supplementation in inhibiting the plaque formation with diminished expression of MCP-1 at the aortic lesion.

Lutein is one of the most widely found carotenoids distributed in fruits and vegetables frequently consumed. Its presence in human tissues is entirely of dietary origin. Distribution of lutein among tissues is similar to other carotenoids but, along with zeaxanthin, they are found selectively at the centre of the retina, being usually referred to as macular pigments. Lutein has no provitamin A activity in man but it displays biological activities that have attracted great attention in relation to human health. Epidemiological studies have shown inconsistent associations between high intake or serum levels of lutein and lower risk for developing cardiovascular disease, several types of cancer, cataracts and age-related maculopathy. Also, lutein supplementation has provided both null and positive results on different biomarkers of oxidative stress although it is effective in increasing macular pigment concentration and in improving visual function in some, but not all, subjects with different eye pathologies. Overall, data suggest that whereas serum levels of lutein have, at present, no predictive, diagnostic or prognostic value in clinical practice, its determination may be very helpful in assessing compliance and efficacy of intervention as well as potential toxicity. In addition, available evidence suggests that a serum lutein concentration between 0·6 and 1·05 μmol/l seems to be a safe, dietary achievable and desirable target potentially associated with beneficial impact on visual function and, possibly, on the development of other chronic diseases. The use of lutein as a biomarker of exposure in clinical practice may provide some rationale for assessing its relationship with human health as well as its potential use within the context of evidence-based medicine.

The shortcomings of the metabolic implications of the current protein requirements model are reviewed, and an alternative model, validated with [1-13C]leucine balance results in human adults, is presented and evaluated in the context of defining protein requirements. The model identifies metabolic demands for amino acids as comprising a small fixed component and a variable adaptive component that is relatively insensitive to acute food or protein intake, but which changes slowly with a sustained change in intake, enabling N equilibrium to be achieved. The model accounts for the apparent low efficiency of utilisation of animal proteins in N balance studies and enables more realistic efficiency values to be measured within an experimental framework that takes account of the adaptive metabolic demand. However, the complex relationship between the adaptive metabolic demand and habitual level and quality of protein intake prevents prediction of protein quality by amino acid scoring, which can markedly underestimate actual values. In contrast to the current model, for fully adapted individuals risk of deficiency (i.e. negative N balance after complete adaptation) will only start to increase when intakes fall below the range of the true minimum requirements, i.e. a value that is currently unknown, but likely to be between 0·40 and 0·50 g/kg per d at the lower end of the reported distribution of requirements. At intakes greater than this with additional metabolic demands varying directly with intake, deficiency is only likely as a short-term response to a change to a lower intake within the adaptive range. Thus, for adults satisfying energy needs on most mixed human diets, intakes will be within the adaptive range, and N equilibrium ceases to be a useful indicator of nutritional adequacy of protein. In the context of prescriptive dietary guidelines it may be expedient to retain current values until the benefits (and any risks) of protein intakes within the adaptive range can be quantified. However, from a diagnostic perspective, indicators other than N balance need to be identified, since maintenance of N balance can no longer be used as a surrogate of adequate protein-related health.

An increased protein content and less digestible protein sources in the diet can induce bad faecal odour. The present study investigated the effect of adding prebiotics to dog diets enriched with animal-derived protein sources on apparent digestibilities and faecal ammonia concentration. In three subsequent periods eight healthy beagle dogs were fed a commercial dog diet that was gradually supplemented by up to 50 % with meat and bone meal (MBM), greaves meal (GM) or poultry meal (PM) respectively. Afterwards, 3 % fructo-oligosaccharides or 3 % isomalto-oligosaccharides were substituted for 3 % of the total diet. Supplementation with animal-derived protein sources did not decrease the apparent N digestibility significantly but oligosaccharides did. On the other hand the bacterial N content (% DM) in the faeces was highest in the oligosaccharide groups followed by the protein-supplemented groups and lowest in the control groups. When the apparent N digestibility was corrected for bacterial N no significant differences were noted anymore except for the GM group where the corrected N digestibility was still lower after oligosaccharide supplementation. The amount of faecal ammonia was significantly increased by supplementing with protein or oligosaccharides in the MBM and GM groups but not in the PM group. When apparent N digestibility is interpreted, a correction for bacterial N should be taken into account, especially when prebiotics are added to the diet. Oligosaccharides did not reduce the faecal ammonia concentrations as expected.

The effect of polyunsaturated fatty acids (PUFA), in particular conjugated linoleic acid (CLA), on Ca and bone metabolism is unclear. In a 2 × 2 factorial design study, forty male 4-week-old rats were fed a control diet containing 70 g added fat (soyabean oil (SBO; n–6 PUFA-rich diet) or menhaden oil–safflower oil (MSO; n−3 PUFA-rich diet))/kg diet with 0 or 10 g CLA/kg for 8 weeks. Ex vivo prostaglandin E2biosynthesis by bone organ culture was significantly higher (P<0·001) in rats consuming SBO compared with MSO, irrespective of CLA. Addition of the CLA treatment to either diet further lowered (P<0·05) ex vivo prostaglandin E2 production. Neither PUFA type nor CLA altered circulating or femoral mRNA levels of osteocalcin (a marker of bone formation) or insulin-like growth factor-I (a mediator of bone metabolism). While urinary pyridinium crosslinks levels (markers of bone resorption) were unaffected by CLA irrespective of PUFA type, they were significantly higher (P<0·05) in rats consuming SBO compared with MSO irrespective of CLA. Net fractional (%) and absolute (mg) Ca absorption were significantly (P<0·01 and P<0·05 respectively) higher in CLA-supplemented than unsupplemented animals fed on the n−3 PUFA-rich diet, whereas CLA had no effect in animals fed the n–6 PUFA-rich diet. There was no effect of CLA supplementation on bone mineral mass. In conclusion, CLA supplementation over 8 weeks appeared to enhance Ca absorption in young growing rats fed an n−3 PUFA-rich diet, but had no measurable effect on bone metabolism or bone mass over this time frame.

Our objective was to determine whether dietary plant proteins such as soya-protein isolate (SPI) and rice-protein isolate (RPI) compared with animal proteins, such as casein, could afford beneficial effects on atherosclerosis development in apolipoprotein E-deficient mice. In experiment 1, male and female mice were fed on a purified diet containing either casein, SPI or RPI for 9 weeks. The en face lesion area in the aorta (P < 0·05) and the lesion size in the aortic root (P < 0·05) in mice fed the casein-based diet were greater than those in the SPI or RPI groups. The plant protein groups had an increased concentration of serum l-arginine (P < 0·05) and NO metabolites (NO2 plus NO3) (P < 0·05) than did the casein group. The inhibitory effect of the plant proteins on the lesion formations was unrelated to gender and total serum cholesterol. In experiment 2, the l-arginine and l-methionine contents were the same in the l-arginine-supplemented casein-based and SPI-based diets, and between the l-methionine-supplemented SPI-based and the casein-based diets. Male mice were fed on the diets for 15 weeks. There were no significant differences in the en face lesion area and the lesion size between the casein group and the l-arginine-supplemented group, although the serum l-arginine (P < 0·05) and NO2 plus NO3 (P < 0·05) concentrations in the supplemented group were higher than those in the casein group. There were no significant effects of l-methionine supplementation on the lesion formations. In experiment 3, male mice were given the casein-based diet or the l-arginine-supplemented casein-based diet together with water or water containing an NO synthesis inhibitor for 9 weeks. When given the casein-based diet, the inhibitor drinking, compared with water drinking, resulted in a reduction of the serum NO2 plus NO3 concentration (P < 0·01) and an increase in the en face lesion area (P < 0·05) and the lesion size (P < 0·01). When given the l-arginine-supplemented diet, the inhibitor drinking, compared with water drinking, resulted in no increase in the lesion area and size. These results demonstrate anti-atherogenic potentials of SPI- as well as RPI-derived proteins, but their l-arginine and l-methionine contents were not sufficient enough to explain the underlying mechanism(s).

To determine the optimum indispensable (I) amino acid (AA) balance in Atlantic salmon (Salmo salar L.) fry, a single protocol established for the pig was adapted. The balance was calculated from the reduction in N gain after replacing about 45% of a single IAA by a mixture of dispensable AA in isonitrogenous diets. We confirmed that the mixture of AA simulating the AA pattern of cod-meal protein and gelatine (46:3, w/w) was used with the same efficiency as cod-meal protein and gelatine. From the deletion experiment an optimum balance between the IAA was derived. Expressed relative to lysine=100, the optimal balance was: arginine 76 (se 0·2), histidine 28 (se 2·2), methionine+cystine 64 (se 1·7), phenylalanine + tyrosine 105 (se 1·6), threonine 51 (se 2·4), tryptophan 14 (se 0·7), valine 59 (se 1·7). No estimates were made for isoleucine and leucine. Expressed as g/16g N, the optimal balance was: arginine 4·0 (se 0·0), histidine 1·5 (se 0·1), lysine 5·3 (se 0·2), methionine+cystine 3·4 (se 0·1), phenylaline+tyrosine 5·6 (se 0·1), threonine 2·7 (se 0·1), tryptophan 0·7 (se 0·0), valine 3·1 (se 0·1). This AA composition is close to that of the Atlantic salmon whole-body, but using it as an estimation of the IAA requirements may lead to an overestimation of the branched-chain AA requirements and an underestimation of aromatic and S-containing AA requirements. The results are discussed in accordance with the key assumptions associated with the model used (broken-line model, IAA efficiencies and maintenance requirements).

A method for measuring unlabelled Fe absorption has been investigated in a pilot study using a simple mathematical model. The metabolism of newly absorbed Fe can be approximated as a single-compartment model with the sampled compartment being the plasma pool. Five female volunteers (aged 30–55 years) were recruited to participate in the pilot study. After a 10 mg oral dose of unlabelled ferrous sulfate, the change in plasma Fe concentration over the following 6 h was used to estimate the quantity of absorbed Fe from the mathematical model. To assess the accuracy of the new technique, a 1 mg oral dose of 57Fe-labelled iron sulfate was given simultaneously with a 225 μg intravenous dose of 58Fe as iron citrate. The plasma appearance of the labelled Fe was used to estimate the absorption of the oral label from the traditional area under the curve method. There was no significant difference (P=0·61) between the geometric mean absorption of the unlabelled (19 (- 1 sd 12, +1 sd 28) %) and the labelled Fe (17 (- 1 sd 10, +1 sd 29) %). These initial results are encouraging, but further work needs to be undertaken with smaller doses, as typically found in meals. The effect of diurnal variation in serum Fe concentration on the estimation of unlabelled Fe absorption needs further assessment.

The objective of the present study was to investigate the effects of dietary macronutrient ratio on energy metabolism and on skeletal muscle mRNA expression of avian uncoupling protein (UCP), thought to be implicated in thermogenesis in birds. Broiler chickens from 2 to 6 weeks of age received one of three isoenergetic diets containing different macronutrient ratios (low-lipid (LL) 30 v. 77 g lipid/kg; low-protein (LP) 125 v. 197 g crude protein (N×6·25)/kg; low-carbohydrate (LC) 440 v. 520 g carbohydrate/kg). LP chickens were characterised by significantly lower body weights and food intakes compared with LL and LC chickens (−47 and −38 % respectively) but similar heat production/kg metabolic body weight, as measured by indirect calorimetry, in the three groups. However, heat production/g food ingested was higher in animals receiving the LP diet (+41 %, P<0·05). These chickens also deposited 57 % less energy as protein (P<0·05) and 33 % more as fat. No significant differences in energy and N balances were detected between LL and LC chickens. The diets with the higher fat contents (i.e. the LP and LC diets) induced slightly but significantly higher relative expressions of avian UCP mRNA in gastrocnemius muscle, measured by reverse transcription–polymerase chain reaction, than the LL diet (88 and 90 v. 78 % glyceraldehyde-3-phosphate dehydrogenase respectively, P<0·05). Our present results are consistent with the recent view that UCP homologues could be involved in the regulation of lipid utilisation as fuel substrate and provide evidence that the macronutrient content of the diet regulates energy metabolism and especially protein and fat deposition.

To investigate the role of the monounsaturated n-9 fatty acids (MUFA) in the lipolytic activity of adipocytes, a study was carried out in which an increase in MUFA was produced in the tissues by two different methods; by the dietary enrichment of oleic acid or by producing an essential fatty acid deficiency syndrome. For this, forty-five male Sprague–Dawley rats were fed with a normal-energy diet and were subdivided into three groups. The diets varied in the type of dietary fat; palmitic acid, olive oil, or soyabean oil+palmitic acid. At the end of the study measurements were taken of weight, plasma leptin, tissue concentration of fatty acids, fat-cell size in the epididymal and the omental adipose tissues, adipocyte lipolytic activity of both tissues after stimulation with adrenaline, and the capacity of insulin to inhibit lipolysis. The baseline and adrenaline-stimulated lipolytic activity were greater and the anti-lipolytic capacity of insulin lower in the animals undergoing an increase in MUFA in the tissues (palmitic-acid and olive-oil diets). The area under the curve of glycerol, used as an indicator of lipolytic activity, was positively correlated with the concentration of MUFA and negatively with polyunsaturated fatty acids in the adipose tissues. It is concluded that an increase in tissue MUFA, however obtained, induces an increase in lipolytic activity.

The cellular uptake of ammonia affects the intracellular pH (pHi) of polar and non-polar cells. A predominant uptake of NH3 and its intra-cellular protonation tend to alkalinise the cytoplasm, whereas a predominant uptake of NH4+ acidifies the cytoplasm by reversing this reaction. Hence, the well-known absorption of ammonia across the rumen epithelium probably causes a change in the pHi. The magnitude and direction of this change in pHi (acid or alkaline) depends on the relative transport rates of NH3 and NH4+. Consequently, the intra-cellular availability of protons will influence the activity of the Na+-H+ exchanger, which could affect transepithelial Na+ transport. The aim of the present study has been to test this possible interaction between ruminal ammonia concentrations and Na+ transport. The term ammonia is used to designate the sum of the protonated (NH4+) and unprotonated (NH3) forms. Isolated ruminal epithelium of sheep was investigated by using the Ussing-chamber technique in vitro. The present results indicate that ammonia inhibits Na+ transport across the rumen epithelium of hay-fed sheep, probably by binding intracellular protons and thus inhibiting Na+-H+ exchange. By contrast, ammonia stimulates Na+ transport in concentrate-fed and urea-fed sheep, which develop an adaptation mechanism in the form of an increased metabolism of ammonia in the rumen mucosa and/or an increased permeability of rumen epithelium to the charged ammonium ion (NH4+). Intracellular dissociation of NH4+ increases the availability of protons, which stimulate Na+ –H+ exchange. This positive effect of ruminal ammonia on Na+ absorption may significantly contribute to the regulation of osmotic pressure of the ruminal fluid, because intraruminal ammonia concentrations up to 40 mmol/l have been reported.