Choline and its derivative betaine are important methyl donors, components of cell membrane phospholipids, or precursors of the neurotransmitter acetylcholine and the gut microbial metabolite trimethylamine-N-oxide. We aimed to investigate trends in dietary intake and food sources of total choline, individual choline forms, and betaine in Chinese adults using data from the China Health and Nutrition Survey (CHNS) 1991-2011. The sample was drawn from urban and rural communities in 12 provinces or autonomous regions with a multistage, random cluster design. Dietary intake was estimated using three consecutive 24-hour dietary recalls in combination with a household food inventory. Linear mixed-effect models were constructed to calculate adjusted mean intake values and 95% confidence intervals (CIs) using R version 4.2.2. A total of 11,188 men and 12,279 women aged 18 years or older were included. Between 1991 and 2011, total choline intake increased from 219.3 (95% CI: 215.1-223.4) mg/d to 269.0 (95% CI: 265.6-272.5) mg/d in men and from 195.6 (95% CI: 191.8-199.4) mg/d to 240.4 (95% CI: 237.4-243.5) mg/d in women (both P-trends <0.001). Phosphatidylcholine was the major form of dietary choline and its contribution to total choline increased from 46.9% in 1991 to 58.8% in 2011. Cereals were the primary food source of total choline before 2000 while eggs had ranked at the top since 2004. Dietary betaine intake was relatively steady over time with a range of 134.0-151.5 mg/d in men (P-trend <0.001) and 111.7-125.3 mg/d in women (P-trend >0.05). In conclusion, Chinese adults experienced a significant increase in dietary intake of choline, particularly phosphatidylcholine during 1991-2011 and animal-derived foods have replaced plant-based foods as the main food sources of choline. Betaine intake remained relatively stable over time. Future efforts should address the health effects of these changes.

Previous animal studies found beneficial effects of choline and betaine on maternal glucose metabolism during pregnancy, but few human studies explored the association between choline or betaine intake and incident gestational diabetes mellitus (GDM). We aimed to explore the correlation of dietary choline or betaine intake with GDM risk among Chinese pregnant women. A total of 168 pregnant women with GDM cases and 375 healthy controls were enrolled at the Seventh People’s Hospital in Shanghai during their GDM screening at 24–28 gestational weeks. A validated semi-quantitative FFQ was used to estimate choline and betaine consumption through face-to-face interviews. An unconditional logistic regression model was adopted to examine OR and 95 % CI. Compared with the controls, those women with GDM incidence were likely to have higher pre-pregnancy BMI, be older, have more parities and have higher plasma TAG and lower plasma HDL-cholesterol. No significant correlation was observed between the consumption of choline or betaine and incident GDM (adjusted OR (95 % CI), 0·77 (0·41, 1·43) for choline; 0·80 (0·42, 1·52) for betaine). However, there was a significant interaction between betaine intake and parity on the risk of GDM (Pfor interaction = 0·01). Among those women with no parity history, there was a significantly inverse correlation between betaine intake and GDM risk (adjusted OR (95 % CI), 0·25 (0·06, 0·81)). These findings indicated that higher dietary betaine intake during pregnancy might be considered a protective factor for GDM among Chinese women with no parity history.

It is inconclusive whether trimethylamine N-oxide (TMAO) and choline and related metabolites, namely trimethylamine (TMA), l-carnitine, betaine and dimethylglycine (DMG), are associated with non-alcoholic fatty liver disease (NAFLD). Our objective was to investigate these potential associations. Additionally, we sought to determine the mediating role of TMAO. In this 1:1 age- and sex-matched case–control study, a total of 150 pairs comprising NAFLD cases and healthy controls were identified. According to the fully adjusted model, after the highest tertile was compared with the lowest tertile, the plasma TMAO concentration (OR = 2·02 (95 % CI 1·04, 3·92); P trend = 0·003), l-carnitine concentration (OR = 1·79 (1·01, 3·17); P trend = 0·020) and DMG concentration (OR = 1·81 (1·00, 3·28); P trend = 0·014) were significantly positively associated with NAFLD incidence. However, a significantly negative association was found for plasma betaine (OR = 0. 50 (0·28, 0·88); P trend = 0·001). The restricted cubic splines model consistently indicated positive dose–response relationships between exposure to TMAO, l-carnitine, and DMG and NAFLD risk, with a negative association being observed for betaine. The corresponding AUC increased significantly from 0·685 (0·626, 0·745) in the traditional risk factor model to 0·769 (0·716, 0·822) when TMAO and its precursors were included (l-carnitine, betaine and choline) (P = 0·032). Mediation analyses revealed that 14·7 and 18·6 % of the excess NAFLD risk associated with l-carnitine and DMG, respectively, was mediated by TMAO (the P values for the mediating effects were 0·021 and 0·036, respectively). These results suggest that a higher concentration of TMAO is associated with increased NAFLD risk among Chinese adults and provide evidence of the possible mediating role of TMAO.

Methyl donor micronutrients might affect muscle strength via DNA methylation. We aimed to evaluate the combined relationship of dietary methyl donor micronutrients containing betaine, choline, methionine, vitamin B12, vitamin B6 and folate on muscle strength. This cross-sectional study was conducted on 267 subjects including 113 men and 154 women. Dietary intake of micronutrients was assessed utilising a validated 168-item semi-quantitative FFQ, and methyl donor micronutrient score (MDMS) was calculated. The muscle strength of the participants was measured using a digital handgrip dynamometer. The association was determined using linear regression analysis. The mean age of participants was 36·8 ± 13·2 years. After taking into account potential confounding variables, there was no significant association between dietary methyl donor micronutrient score (MDMS) and the mean left-hand muscle strength (β: 0·07, se: 0·05, P = 0·07); however, the changes were significant in the mean right-hand muscle strength (β: 0·09, se: 0·04, P = 0·03). There was also a significant positive relationship between mean muscle strength and methyl donors’ intake after fully adjusting for potential confounders (β: 0·08, se: 0·04, P = 0·04). In conclusion, our findings revealed that higher dietary methyl donor micronutrient consumption is associated with enhanced muscle strength. As a result, advice on a higher intake of methyl donor-rich foods including grains, nuts, dairy products and seafood might be recommended by dietitians as a general guideline to adhere to. Additional prospective studies are needed to confirm the findings.

One-carbon nutrients play an important role in epigenetic mechanisms and cellular methylation reactions. Inadequate intake of these nutrients is linked to metabolic perturbations, yet the current intake levels of these nutrients have rarely been studied in Asia. This cross-sectional study surveyed the usual dietary intake of one-carbon nutrients (folate, choline and vitamins B2, B6 and B12) among Thai university students aged 19–30 years (n 246). Socioeconomic background, health information, anthropometric data and 24-h dietary recall data were collected. The long-term usual intake was estimated using the multiple-source method. The average usual intake levels for men and women were (mean ± sd) 1⋅85 ± 0⋅95 and 2⋅42 ± 8⋅7 mg/d of vitamin B2, 1⋅96 ± 1⋅0 and 2⋅49 ± 8⋅7 mg/d of vitamin B6, 6⋅20 ± 9⋅5 and 6⋅28 ± 12 μg/d of vitamin B12, 195 ± 154 and 155 ± 101 μg dietary folate equivalent/d of folate, 418 ± 191 and 337 ± 164 mg/d of choline, respectively. Effect modification by sex was observed for vitamin B2 (P-interaction = 0⋅002) and choline (P-interaction = 0⋅02), where every 1 mg increase in vitamin B2 and 100 mg increase in choline intake were associated with a 2⋅07 (P = 0⋅01) and 0⋅81 kg/m2 (P = 0⋅04) lower BMI, respectively, in men. The study results suggest that Thai young adults meet the recommended levels for vitamins B2, B6 and B12. The majority of participants had inadequate folate intake and did not achieve recommended intake levels for choline. The study was approved by the Ethics Committee at the Faculty of Medicine, Chiang Mai University. This trial was registered at www.thaiclinicaltrials.gov (TCTR20210420007).

The transfer of one-carbon units between molecules in metabolic pathways is essential for maintaining cellular homeostasis, but little is known about whether the circulating concentrations of metabolites involved in the one-carbon metabolism are affected by the prandial status. Epidemiological studies do not always consistently use fasting or non-fasting blood samples or may lack information on the prandial status of the study participants. Therefore, the main aim of the present study was to investigate the effects of a light breakfast on serum concentrations of selected metabolites and B-vitamins related to the one-carbon metabolism; i.e. the methionine-homocysteine cycle, the folate cycle, the choline oxidation pathway and the transsulfuration pathway. Sixty-three healthy adults (thirty-six women) with BMI ≥ 27 kg/m2 were included in the study. Blood was collected in the fasting state and 60 and 120 min after intake of a standardised breakfast consisting of white bread, margarine, white cheese, strawberry jam and orange juice (2218 kJ). The meal contained low amounts of choline, betaine, serine and vitamins B2, B3, B6, B9 and B12. Serum concentrations of total homocysteine, total cysteine, flavin mononucleotide, nicotinamide and pyridoxal 5’-phosphate were significantly decreased, and concentrations of choline, betaine, dimethylglycine, sarcosine, cystathionine and folate were significantly increased following breakfast intake (P < 0·05). Our findings demonstrate that the intake of a light breakfast with low nutrient content affected serum concentrations of several metabolites and B-vitamins related to the one-carbon metabolism.

The increasing prevalence of non-alcoholic fatty liver disease (NAFLD) poses a growing challenge in terms of its prevention and treatment. The ‘multiple hits’ hypothesis of multiple insults, such as dietary fat intake, de novo lipogenesis, insulin resistance, oxidative stress, mitochondrial dysfunction, gut dysbiosis and hepatic inflammation, can provide a more accurate explanation of the pathogenesis of NAFLD. Betaine plays important roles in regulating the genes associated with NAFLD through anti-inflammatory effects, increased free fatty oxidation, anti-lipogenic effects and improved insulin resistance and mitochondrial function; however, the mechanism of betaine remains elusive.

Nonalcoholic fatty liver disease (NAFLD) represents a hepatic manifestation of metabolic syndrome. The aim of this study was to examine the effect of betaine on ultrastructural changes in the mouse liver with methionine- and choline-deficient (MCD) diet-induced NAFLD. Male C57BL/6 mice were divided into groups: Control—fed with standard chow, BET—standard chow supplemented with betaine (1.5% w/v drinking water), MCD—fed with MCD diet, and MCD + BET—MCD diet with betaine supplementation for 6 weeks. Liver samples were taken for pathohistology and transmission electron microscopy. The MCD diet-induced steatosis, inflammation, and balloon-altered hepatocytes were alleviated by betaine. MCD diet induced an increase in mitochondrial size versus the control group (p < 0.01), which was decreased in the betaine-treated group. In the MCD diet-fed group, the total mitochondrial count decreased versus the control group (p < 0.01), while it increased in the MCD + BET group versus MCD (p < 0.01). Electron microscopy showed an increase in the number of autophagosomes in the MCD and MCD + BET group versus control, and a significant difference in autophagosomes number was detected in the MCD + BET group by comparison with the MCD diet-treated group (p < 0.05). Betaine decreases the number of enlarged mitochondria, alleviates steatosis, and increases the number of autophagosomes in the liver of mice with NAFLD.

This study evaluated the effects of rumen-protected folic acid (RPFA) and betaine (BT) on growth performance, nutrient digestion and blood metabolites in bulls. Forty-eight Angus bulls were blocked by body weight and randomly assigned to four treatments in a 2 × 2 factorial design. BT of 0 or 0·6 g/kg DM was supplemented to diet without or with the addition of 6 mg/kg DM of folic acid from RPFA, respectively. Average daily gain increased by 25·2 and 6·29 % for addition of BT without RPFA and with RPFA, respectively. Digestibility and ruminal total volatile fatty acids of neutral-detergent fibre and acid-detergent fibre increased, feed conversion ratio and blood folate decreased with the addition of BT without RPFA, but these parameters were unchanged with BT addition in diet with RPFA. Digestibility of DM, organic matter and crude protein as well as acetate:propionate ratio increased with RPFA or BT addition. Ruminal ammonia-N decreased with RPFA addition. Activity of carboxymethyl cellulase, cellobiase, xylanase, pectinase and protease as well as population of total bacteria, protozoa, Fibrobacter succinogenes and Ruminobacter amylophilus increased with RPFA or BT addition. Laccase activity and total fungi, Ruminococcus flavefaciens and Prevotella ruminicola population increased with RPFA addition, whereas Ruminococcus albus population increased with BT addition. Blood glucose, total protein, albumin, growth hormone and insulin-like growth factor-1 increased with RPFA addition. Addition of RPFA or BT decreased blood homocysteine. The results indicated that addition of BT stimulated growth and nutrient digestion in bulls only when RPFA was not supplemented.

There is a growing research interest in determining whether changes in the global status of DNA methylation are related to the environment, in particular, to one-carbon metabolism. So, our aim was to investigate the effect of dietary methyl-group donor intake (methionine, folate, choline, betaine, vitamins B2, B6 and B12), biomarkers (total folate, unmetabolised folic acid (FA), 5-methyltetrahydrofolate, homocysteine, vitamins B6 and B12 concentrations) and genetic variants (polymorphisms involved in one-carbon metabolism) on global DNA methylation in a population exposed to mandatory flour fortification with FA. A cross-sectional study of health and living conditions was conducted among a representative sample of residents in São Paulo, Brazil. The mean of global DNA methylation was lower in young people than in adults and the elderly (P = 0·049). No differences between genotypes of polymorphism and global DNA methylation mean were identified. We observed that the increase in betaine intake led to an absolute change in percentage of DNA methylation (β = 0·0005, P = 0·024) using multiple regression. Betaine intake alone was associated with an absolute change in percentage of global DNA methylation. The study did not find an association between global DNA methylation and folate status even in a population exposed to mandatory flour fortification with FA.

Maternal one-carbon metabolism during pregnancy is crucial for fetal development and programming by DNA methylation. However, evidence on one-carbon biomarkers other than folate is lacking. We, therefore, investigated whether maternal plasma methyl donors, that is, choline, betaine and methionine, are associated with birth outcomes. Blood samples were obtained from 115 women during gestation (median 26·3 weeks, 90 % range 22·7–33·0 weeks). Plasma choline, betaine, methionine and dimethylglycine were measured using HPLC-tandem MS. Multivariate linear and logistic regression models were used to estimate the association between plasma biomarkers and birth weight, birth length, the risk of small-for-gestational-age and large-for-gestational-age (LGA). Higher level of maternal betaine was associated with lower birth weight (–130·3 (95 % CI –244·8, –15·9) per 1 sd increment for log-transformed betaine). Higher maternal methionine was associated with lower risk of LGA, and adjusted OR, with 95 % CI for 1 sd increase in methionine concentration was 0·44 (95 % CI 0·21, 0·89). Stratified analyses according to infant sex or maternal plasma homocysteine status showed that reduction in birth weight in relation to maternal betaine was only limited to male infants or to who had higher maternal homocysteine status (≥5·1 µmol/l). Higher maternal betaine status was associated with reduced birth weight. Maternal methionine was inversely associated with LGA risk. These findings are needed to be replicated in future larger studies.

The effects of betaine on hepatocytes chromatin architecture changes were examined by using fractal and gray-level co-occurrence matrix (GLCM) analysis in methionine/choline-deficient (MCD) diet-induced, nonalcoholic fatty liver disease (NAFLD). Male C57BL/6 mice were divided into groups: (1) Control: standard diet; (2) BET: standard diet and betaine supplementation through drinking water (solution 1.5%); (3) MCD group: MCD diet for 6 weeks; (4) MCD+BET: fed with MCD diet + betaine for 6 weeks. Liver tissue was collected for histopathology, immunohistochemistry, and determination of fractal dimension and GLCM parameters. MCD diet induced diffuse micro- and macrovesicular steatosis accompanied with increased Ki67-positive hepatocyte nuclei. Steatosis and Ki67 immunopositivity were less prominent in the MCD+BET group compared with the MCD group. Angular second moment (ASM) and inverse difference moment (IDM) (textural homogeneity markers) were significantly increased in the MCD+BET group versus the MCD group (p<0.001), even though no difference between the MCD and the control group was evident. Heterogeneity parameters, contrast, and correlation were significantly increased in the MCD group versus the control (p<0.001). On the other hand, betaine treatment significantly reduced correlation, contrast, and entropy compared with the MCD group (p<0.001). Betaine attenuated MCD diet-induced NAFLD by reducing fat accumulation and inhibiting hepatocyte proliferation. Betaine supplementation increased nuclear homogeneity and chromatin complexity with reduction of entropy, contrast, and correlation.

The isolated or combined effects of betaine and arginine supplementation of reduced protein diets (RPD) on fat content, fatty acid composition and mRNA levels of genes controlling lipid metabolism in pig m. longissimus lumborum and subcutaneous adipose tissue (SAT) were assessed. The experiment was performed on forty intact male pigs (Duroc×Large White×Landrace cross-breed) with initial and final live weights of 60 and 93 kg, respectively. Pigs were randomly assigned to one of the following five diets (n 8): 16·0 % of crude protein (control), 13·0 % of crude protein (RPD), RPD supplemented with 0·33 % of betaine, RPD supplemented with 1·5 % of arginine and RPD supplemented with 0·33 % of betaine and 1·5 % of arginine. Data confirmed that RPD increase intramuscular fat (IMF) content and total fat content in SAT. The increased total fat content in SAT was accompanied by higher GLUT type 4, lipoprotein lipase and stearoyl-CoA desaturase mRNA expression levels. In addition, the supplementation of RPD with betaine and/or arginine did not affect either IMF or total fat in SAT. However, dietary betaine supplementation slightly affected fatty acid composition in both muscle and SAT. This effect was associated with an increase of carnitine O-acetyltransferase mRNA levels in SAT but not in muscle, which suggests that betaine might be involved in the differential regulation of some key genes of lipid metabolism in pig muscle and SAT. Although the arginine-supplemented diet decreased the mRNA expression level of PPARG in muscle and SAT, it did not influence fat content or fatty acid composition in any of these pig tissues.

To assess the effects of betaine on hepatic lipid accumulation and investigate the underlying mechanism, thirty-two male Sprague–Dawley rats weighing 100 (sd 2·50) g were divided into four groups, and started on one of four treatments: basal diet, basal diet with betaine administration, high-fat diet and high-fat diet with betaine administration. The results showed that no significant difference of body weight was found among experimental groups. Compared with high-fat diet-fed rats, a betaine supplementation decreased (P< 0·05) hepatic TAG accumulation induced by high-fat diet, which was also supported by hepatic histology results. Additionally, hepatic betaine–homocysteine methyltransferase activity as well as its mRNA abundance and lecithin level were found increased (P< 0·05) by betaine supplementation in both basal diet-fed rats and high-fat diet-fed rats. Betaine administration in high-fat diet-fed rats exhibited a higher (P< 0·05) activity of hepatic carnitine palmitoyltransferase 1 (CPT1) compared with high-fat diet-fed rats. High-fat diet inhibited (P< 0·05) the gene expression of hepatic PPARα and CPT1. However, betaine administration in high-fat diet-fed rats elevated (P< 0·05) the gene expression of PPARα and CPT1. Moreover, concentration, gene and protein expressions of hepatic fibroblast growth factor 21 (FGF21) were increased (P< 0·05) in response to betaine administration in high-fat diet group; meanwhile the gene expression of hepatic AMP-activated protein kinase was increased (P< 0·05) as well. The results suggest that betaine administration enhanced hepatic lipid export and fatty acid oxidation in high-fat diet-fed rats, thus effectively alleviating fat accumulation in the liver.

The bran and particularly the aleurone fraction of wheat are high in betaine and other physiological methyl donors, which may exert beneficial physiological effects. We conducted two randomised, controlled, cross-over postprandial studies to assess and compare plasma betaine and other methyl donor-related responses following the consumption of minimally processed bran and aleurone fractions (study A) and aleurone bread (study B). For both studies, standard pharmacokinetic parameters were derived for betaine, choline, folate, dimethylglycine (DMG), total homocysteine and methionine from plasma samples taken at 0, 0·5, 1, 2 and 3 h. In study A (n 14), plasma betaine concentrations were significantly and substantially elevated from 0·5 to 3 h following the consumption of both bran and aleurone compared with the control; however, aleurone gave significantly higher responses than bran. Small, but significant, increases were also observed in DMG measures; however, no significant responses were observed in other analytes. In study B (n 13), plasma betaine concentrations were significantly and substantially higher following consumption of the aleurone bread compared with the control bread; small, but significant, increases were also observed in DMG and folate measures in response to consumption of the aleurone bread; however, no significant responses were observed in other analytes. Peak plasma betaine concentrations, which were 1·7–1·8 times the baseline levels, were attained earlier following the consumption of minimally processed aleurone compared with the aleurone bread (time taken to reach peak concentration 1·2 v. 2·1 h). These results showed that the consumption of minimally processed wheat bran, and particularly the aleurone fraction, yielded substantial postprandial increases in plasma betaine concentrations. Furthermore, these effects appear to be maintained when aleurone was incorporated into bread.

The objectives of this study were to determine the effects of conjugated linoleic acid (CLA) or betaine on the growth performance, carcass characteristics and fatty acid composition in backfat and belly fat of pigs fed distillers dried grains with solubles (DDGS). Thirty-two (60±2 kg) crossbred barrows (Duroc×Landrace×Yorkshine) were assigned to one of four diets randomly: (1) the control diet containing no corn DDGS (control group); (2) the diet containing 30% corn DDGS (DDGS-fed group); (3) the diet containing 30% corn DDGS and 10 g/kg CLA (CLA-fed group); (4) the diet containing 30% corn DDGS and 1 g/kg BET (BET-fed group). The pigs fed DDGS showed that the percentages of C18:2, polyunsaturated fatty acid (PUFA) and iodine value (IV) increased, while C18:1, saturated fatty acid (SFA) and monounsaturated fatty acid (MUFA) decreased. Pigs fed the DDGS+CLA or DDGS+betaine diets showed the increased percentage of SFA, and the decreased percentage of C18:2, PUFA and IV. In conclusion, results confirmed that the diets containing 30% DDGS had no detrimental effects on growth performance, but increased the percentage of PUFA and IV and decreased the percentage of SFA and MUFA in the backfat and belly fat. However, supplementation with CLA or BET can part reverse these effects on carcass fat in finishing pigs.

To elucidate the effects of maternal dietary betaine supplementation on hepatic expression of cholesterol metabolic genes in newborn piglets and the involved epigenetic mechanisms, we fed gestational sows with control or betaine-supplemented diets (3 g/kg) throughout pregnancy. Neonatal piglets born to betaine-supplemented sows had higher serum methionine concentration and hepatic content of betaine, which was associated with significantly up-regulated hepatic expression of glycine N-methyltransferase. Prenatal betaine exposure increased hepatic cholesterol content and modified the hepatic expression of cholesterol metabolic genes in neonatal piglets. Sterol regulatory element-binding protein 2 was down-regulated at both mRNA and protein levels, while 3-hydroxy-3-methylglutaryl CoA reductase (HMGCR) was down-regulated at the mRNA level, but up-regulated at the protein level, in betaine-exposed piglets. The transcriptional repression of HMGCR was associated with CpG island hypermethylation and higher repressive histone mark H3K27me3 (histone H3 lysine 27 trimethylation) on the promoter, whereas increased HMGCR protein content was associated with significantly decreased expression of miR-497. Furthermore, LDL receptor was significantly down-regulated at both mRNA and protein levels in the liver of betaine-exposed piglets, which was associated with promoter CpG hypermethylation. In addition, the expression of cholesterol-27α-hydroxylase (CYP27α1) was up-regulated at both mRNA and protein levels, while the expression of cholesterol-7α-hydroxylase (CYP7α1) was increased at the mRNA level, but unchanged at the protein level associated with increased expression of miR-181. These results indicate that maternal betaine supplementation increases hepatic cholesterol content in neonatal piglets through epigenetic regulations of cholesterol metabolic genes, which involve alterations in DNA and histone methylation and in the expression of microRNA targeting these genes.

Guanidinoacetic acid (GAA) is the natural biosynthetic precursor of creatine, in a metabolic reaction that requires only a methyl group transfer. The use of GAA as a food additive for restoring creatine load in human tissues is rather unexplored and data on efficacy and safety are limited. In particular, an increase in serum homocysteine after GAA administration can be regarded as critical and should be prevented. The present study evaluated the effects of orally administered GAA with and without methyl group donors on serum and urine creatine concentrations, and the occurrence of adverse events during an intervention in healthy human subjects. A total of twenty male and female volunteers were randomised in a double-blind design to receive either GAA (2·4 g/d) or GAA with methyl donors (2·4 g/d of GAA and 1·6 g/d of betaine HCl, 5 μg/d of vitamin B12, 10 mg/d of vitamin B6 and 600 μg/d of folic acid) by oral administration for 8 weeks. Serum and urine creatine increased significantly from before to after administration in both groups (P< 0·001). The proportion of participants who reported minor adverse events was 33·3 % in the GAA group, and 10·0 % in the GAA with methyl donors group (P= 0·30). Hyperhomocysteinaemia was found in 55·6 % of participants supplemented with GAA, while no participant experienced hyperhomocysteinaemia in the group supplemented with GAA and methyl donors (P= 0·01). In summary, both interventions strongly influenced creatine metabolism, resulting in a significant increase in fasting serum creatine. The concomitant supplementation of methyl donors along with GAA largely precluded the elevation of serum homocysteine caused by GAA administration alone.

Obesity is characterised by a state of chronic low-grade inflammation and the elevated circulating and tissue levels of inflammatory markers, including inflammation-related adipokines, released from white adipose tissue. The expression and release of these adipokines generally rises as the adipose tissue expands and hypoxic conditions start to develop within the tissue. Here, the effect of betaine, a trimethylglycine having a biological role as an osmolyte and a methyl donor, on the expression of inflammation-related markers was tested in human adipocytes under hypoxia. Differentiated adipocytes were cultivated under low (1 %) oxygen tension for 8–20 h. The expression of different adipokines, including IL-6, leptin, PPARγ, TNF-α and adiponectin, was measured by quantitative PCR by determining the relative mRNA level from the adipocytes. Hypoxia, in general, led to a decrease in the expression of PPARγ mRNA in human adipocytes, whereas the expression levels of leptin and IL-6 mRNA were substantially increased by hypoxia. The cultivation of adipocytes under hypoxia also led to a reduction in the expression of TNF-α mRNA. The results showed that hypoxia increased the relative quantification of leptin gene transcription, and that betaine (250 μmol/l) reduced this effect, caused by low oxygen conditions. Under hypoxia, betaine also reduced the mRNA level of the pro-inflammatory markers IL-6 and TNF-α. These results demonstrate that the extensive changes in the expression of inflammation-related adipokines in human adipocytes caused by hypoxia can be diminished by the presence of physiologically relevant concentrations of betaine.

To determine possible mechanisms of action that might explain the nutrient partitioning effect of betaine and conjugated linoleic acid (CLA) in Iberian pigs and to address potential adverse effects, twenty gilts were restrictively fed from 20 to 50 kg BW Control, 0.5% betaine, 1% CLA or 0.5% betaine + 1% CLA diets. Serum hormones and metabolites profile were determined at 30 kg BW and an oral glucose test was performed before slaughter. Pigs were slaughtered at 50 kg BW and livers were obtained for chemical and histological analysis. Decreased serum urea in pigs fed betaine and betaine + CLA diets (11%; P = 0.0001) indicated a more efficient N utilization. The increase in serum triacylglycerol (58% and 28%, respectively; P = 0.0098) indicated that CLA and betaine + CLA could have reduced adipose tissue triacylglycerol synthesis from preformed fatty acids. Serum glucose, low-density lipoprotein (LDL) cholesterol and non-esterified fatty acids were unaffected. CLA and betaine + CLA altered serum lipids profile, although liver of pigs fed CLA diet presented no histopathological changes and triglyceride content was not different from Control pigs. Compared with controls, serum growth hormone decreased (20% to 23%; P = 0.0209) for all treatments. Although serum insulin increased in CLA, and especially in betaine + CLA pigs (28% and 83%; P = 0.0001), indices of insulin resistance were unaffected. In conclusion, CLA, and especially betaine + CLA, induced changes in biochemical parameters and hormones that may partially explain a nutrient partitioning effect in young pigs. Nevertheless, they exhibited weak, although detrimental, effects on blood lipids. Moreover, although livers were chemically and histologically normal, pigs fed CLA diet challenged with a glucose load had higher serum glucose than controls.