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Background: The generation of the active form of vitamin B-6, pyridoxal 5'-phosphate (PLP), in tissues is dependent upon riboflavin as flavin mononucleotide, but whether this interaction is important for maintaining vitamin B-6 status is unclear., Objective: To investigate vitamin B-6 and riboflavin status, their metabolic interaction, and relationship with methylenetetrahydrofolate reductase (MTHFR) genotype in adulthood., Methods: Data from 5612 adults aged 18-102 y were drawn from the Irish National Adult Nutrition Survey (NANS; population-based sample) and the Trinity-Ulster Department of Agriculture (TUDA) and Genovit cohorts (volunteer samples). Plasma PLP and erythrocyte glutathione reductase activation coefficient (EGRac), as a functional indicator of riboflavin, were determined., Results: Older (≥65 y) compared with younger (<65 y) adults had significantly lower PLP concentrations (P < 0.001). A stepwise decrease in plasma PLP was observed across riboflavin categories, from optimal (EGRac ≤1.26), to suboptimal (EGRac: 1.27-1.39), to deficient (EGRac ≥1.40) status, an effect most pronounced in older adults (mean ± SEM: 76.4 ± 0.9 vs 65.0 ± 1.1 vs 55.4 ± 1.2 nmol/L; P < 0.001). In individuals with the variant MTHFR 677TT genotype combined with riboflavin deficiency, compared with non-TT (CC/CT) genotype participants with sufficient riboflavin, we observed PLP concentrations of 52.1 ± 2.9 compared with 76.8 ±0.7 nmol/L (P < 0.001). In participants with available dietary data (i.e., NANS cohort, n = 936), PLP was associated with vitamin B-6 intake (nonstandardized regression coefficient β: 2.49; 95% CI 1.75, 3.24; P < 0.001), supplement use (β: 81.72; 95% CI: 66.01, 97.43; P < 0.001), fortified food (β: 12.49; 95% CI: 2.08, 22.91; P = 0.019), and EGRac (β: -65.81; 95% CI: -99.08, -32.54; P < 0.001), along with BMI (β: -1.81; 95% CI: -3.31, -0.30; P = 0.019)., Conclusions: These results are consistent with the known metabolic dependency of PLP on flavin mononucleotide (FMN) and suggest that riboflavin may be the limiting nutrient for maintaining vitamin B-6 status, particularly in individuals with the MTHFR 677TT genotype. Randomized trials are necessary to investigate the PLP response to riboflavin intervention within the dietary range. The TUDA study and the NANS are registered at www.ClinicalTrials.gov as NCT02664584 (27 January 2016) and NCT03374748 (15 December 2017), respectively.Clinical Trial Registry details: Trinity-Ulster-Department of Agriculture (TUDA) study, ClinicalTrials.gov no. NCT02664584 (January 27th 2016); National Adult Nutrition Survey (NANS), ClinicalTrials.gov no. NCT03374748 (December 15th 2017)., (© The Author(s) 2022. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Background: Early applications of metabolomics in nutrition and health research identified associations between dietary patterns and metabolomic profiles. Twin studies show that diet-related phenotypes and diet-associated metabolites are influenced by genes. However, studies have not examined whether diet-metabolite associations are explained by genetic or environmental factors and whether these associations are reproducible over multiple time points., Objective: This research aims to examine the genetic and environmental factors influencing covariation in diet-metabolite associations that are reproducible over time in healthy twins., Methods: The UCD Twin Study is a semi-longitudinal classic twin study that collected repeated dietary, anthropometric, and urinary data over 2 months. Correlation analysis identified associations between diet quality measured using the Healthy Eating Index (HEI) and urinary metabolomic profiles at 3 time points. Diet-associated metabolites were examined using linear regression to identify those significantly influenced by familial factors between twins and those significantly influenced by unique factors. Cholesky decomposition modeling quantified the genetic and environmental path coefficients through associated dietary components onto the metabolites., Results: The HEI was associated with 14 urinary metabolites across 3 metabolomic profiles (r: ±0.15-0.49). For 8 diet-metabolite associations, genetic or shared environmental factors influencing HEI component scores significantly influenced variation in metabolites (β: 0.40-0.52). A significant relation was observed between dietary intakes of whole grain and acetoacetate (β: -0.50, P < 0.001) and β-hydroxybutyrate (β: -0.46, P < 0.001), as well as intakes of saturated fat and acetoacetate (β: 0.47, P < 0.001) and β-hydroxybutyrate (β: 0.52, P < 0.001). For these diet-metabolite associations a common shared environmental factor explained 66-69% of variance in the metabolites., Conclusions: This study shows that diet-metabolite associations are reproducible in 3 urinary metabolomic profiles. Components of the HEI covary with metabolites, and covariation is largely due to the shared environment., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Background: Improved assessment of meat intake with the use of metabolomics-derived markers can provide objective data and could be helpful in clarifying proposed associations between meat intake and health., Objective: The objective of this study was to identify novel markers of chicken intake using a metabolomics approach and use markers to determine intake in an independent cohort., Methods: Ten participants [age: 62 y; body mass index (in kg/m2): 28.25] in the NutriTech food intake study consumed increasing amounts of chicken, from 88 to 290 g/d, in a 3-wk span. Urine and blood samples were analyzed by nuclear magnetic resonance and mass spectrometry, respectively. A multivariate data analysis was performed to identify markers associated with chicken intake. A calibration curve was built based on dose-response association using NutriTech data. A Bland-Altman analysis evaluated the agreement between reported and calculated chicken intake in a National Adult Nutrition Survey cohort., Results: Multivariate data analysis of postprandial and fasting urine samples collected in participants in the NutriTech study revealed good discrimination between high (290 g/d) and low (88 g/d) chicken intakes. Urinary metabolite profiles showed differences in metabolite levels between low and high chicken intakes. Examining metabolite profiles revealed that guanidoacetate increased from 1.47 to 3.66 mmol/L following increasing chicken intakes from 88 to 290 g/d (P < 0.01). Using a calibration curve developed from the NutriTech study, chicken intake was calculated through the use of data from the National Adult Nutrition Survey, in which consumers of chicken had a higher guanidoacetate excretion (0.70 mmol/L) than did nonconsumers (0.47 mmol/L; P < 0.01). A Bland-Altman analysis revealed good agreement between reported and calculated intakes, with a bias of -30.2 g/d. Plasma metabolite analysis demonstrated that 3-methylhistidine was a more suitable indicator of chicken intake than 1-methylhistidine., Conclusions: Guanidoacetate was successfully identified and confirmed as a marker of chicken intake, and its measurement in fasting urine samples could be used to determine chicken intake in a free-living population. This trial was registered at clinicaltrials.gov as NCT01684917., (© 2017 American Society for Nutrition.)

The NOVA classification of foods proposes 4 categories: unprocessed or minimally processed foods, processed culinary ingredients, processed foods, and ultra-processed foods and drinks (UPFDs). It is argued that the latter relies heavily on modifications to foods, resulting in enhanced amounts of salt, added sugar, and fat as well as the use of additives in an attempt to make this food category highly palatable. It further argues that controlling food processing, rather than examining nutrients, should be foremost in shaping nutrition policy. This commentary challenges many of the basic arguments of using the NOVA food classification system to examine the link between food and health. We believe that there is no evidence to uphold the view that UPFDs give rise to hyperpalatable foods associated with a quasi-addictive effect and that the prevailing European Union and US data fail to uphold the assertion that UPFDs, which dominate energy intake, give rise to dietary patterns that are low in micronutrients. With regard to the use of the NOVA food classification in the development of food-based dietary guidelines, we show that the very broad definition of UPFDs makes this impossible. Finally, the available evidence does not support the view that the globalization of food is the driver of increased intakes of UPFDs in low- to middle-income countries but rather that this is driven by small indigenous companies. On balance, therefore, there seems to be little advantage from the use of the NOVA classification compared with the current epidemiologic approach, which relies on the linkage of nutrient intakes to chronic disease with subsequent identification of foods that merit consideration in public health nutrition strategies., (© 2017 American Society for Nutrition.)

Background: There has been limited evidence about whether genotype-tailored advice provides extra benefits in reducing obesity-related traits compared with the benefits of conventional one-size-fits-all advice. Objective: We determined whether the disclosure of information on fat-mass and obesity-associated ( FTO ) genotype risk had a greater effect on a reduction of obesity-related traits in risk carriers than in nonrisk carriers across different levels of personalized nutrition. Design: A total of 683 participants (women: 51%; age range: 18-73 y) from the Food4Me randomized controlled trial were included in this analysis. Participants were randomly assigned to 4 intervention arms as follows: level 0, control group; level 1, dietary group; level 2, phenotype group; and level 3, genetic group. FTO (single nucleotide polymorphism rs9939609) was genotyped at baseline in all participants, but only subjects who were randomly assigned to level 3 were informed about their genotypes. Level 3 participants were stratified into risk carriers (AA/AT) and nonrisk carriers (TT) of the FTO gene for analyses. Height, weight, and waist circumference (WC) were self-measured and reported at baseline and months 3 and 6. Results: Changes in adiposity markers were greater in participants who were informed that they carried the FTO risk allele (level 3 AT/AA carriers) than in the nonpersonalized group (level 0) but not in the other personalized groups (level 1 and 2). Mean reductions in weight and WC at month 6 were greater for FTO risk carriers than for noncarriers in the level 3 group [-2.28 kg (95% CI: -3.06, -1.48 kg) compared with -1.99 kg (-2.19, -0.19 kg), respectively ( P = 0.037); and -4.34 cm (-5.63, -3.08 cm) compared with -1.99 cm (-4.04, -0.05 cm), respectively, ( P = 0.048)]. Conclusions: There are greater body weight and WC reductions in risk carriers than in nonrisk carriers of the FTO gene. This trial was registered at clinicaltrials.gov as NCT01530139., (© 2017 American Society for Nutrition.)

Background: The apolipoprotein E (APOE) risk allele (ɛ4) is associated with higher total cholesterol (TC), amplified response to saturated fatty acid (SFA) reduction, and increased cardiovascular disease. Although knowledge of gene risk may enhance dietary change, it is unclear whether ɛ4 carriers would benefit from gene-based personalized nutrition (PN)., Objectives: The aims of this study were to 1) investigate interactions between APOE genotype and habitual dietary fat intake and modulations of fat intake on metabolic outcomes; 2) determine whether gene-based PN results in greater dietary change than do standard dietary advice (level 0) and nongene-based PN (levels 1-2); and 3) assess the impact of knowledge of APOE risk (risk: E4+, nonrisk: E4-) on dietary change after gene-based PN (level 3)., Design: Individuals (n = 1466) recruited into the Food4Me pan-European PN dietary intervention study were randomly assigned to 4 treatment arms and genotyped for APOE (rs429358 and rs7412). Diet and dried blood spot TC and ω-3 (n-3) index were determined at baseline and after a 6-mo intervention. Data were analyzed with the use of adjusted general linear models., Results: Significantly higher TC concentrations were observed in E4+ participants than in E4- (P < 0.05). Although there were no significant differences in APOE response to gene-based PN (E4+ compared with E4-), both groups had a greater reduction in SFA (percentage of total energy) intake than at level 0 (mean ± SD: E4+, -0.72% ± 0.35% compared with -1.95% ± 0.45%, P = 0.035; E4-, -0.31% ± 0.20% compared with -1.68% ± 0.35%, P = 0.029). Gene-based PN was associated with a smaller reduction in SFA intake than in nongene-based PN (level 2) for E4- participants (-1.68% ± 0.35% compared with -2.56% ± 0.27%, P = 0.025)., Conclusions: The APOE ɛ4 allele was associated with higher TC. Although gene-based PN targeted to APOE was more effective in reducing SFA intake than standard dietary advice, there was no difference between APOE "risk" and "nonrisk" groups. Furthermore, disclosure of APOE nonrisk may have weakened dietary response to PN. This trial was registered at clinicaltrials.gov as NCT01530139., (© 2016 American Society for Nutrition.)

Background: Studies examining vitamin E intake and the percentage of the population meeting dietary guidelines do not distinguish between natural (RRR-α-tocopherol) and synthetic (all-rac-α-tocopherol) intake, even though these different isomeric forms differ in bioactivity., Objective: This study aimed to determine the effect of RRR-α-tocopherol vs. all-rac-α-tocopherol intake on the percentage of the population meeting the vitamin E recommendation and on plasma α-tocopherol stereoisomer distribution., Methods: With the use of data from the Irish National Adult Nutrition Survey (NANS), this study examined the percentage of the Irish population meeting the European Union (EU) RDA for vitamin E of 12 mg/d, correcting for a bioactivity difference in all-rac- vs. RRR-α-tocopherol, where 1 mg of all-rac-α-tocopherol is considered to be equivalent to 1:1.36 (0.74) mg in the EU RDA. In a subcohort of supplement users and nonusers, plasma α- and γ-tocopherol concentrations and α-tocopherol stereoisomer distribution were measured. Receiver operating characteristic (ROC) curve analysis was conducted to determine ability to discriminate supplement user types., Results: Analysis of the NANS showed that 100% of participants still met the recommended intake of 12 mg/d, after all-rac-α-tocopherol intake was corrected for α-tocopherol equivalent bioactivity. In the subcohort analysis, the percentage of plasma RRR-α-tocopherol was significantly lower in high all-rac-α-tocopherol supplement (>11 mg/d) users (82%) compared with nonusers and with high RRR-α-tocopherol supplement (>35 mg/d) users (91% and 93% respectively, P < 0.01). High RRR-α-tocopherol supplement users had a significantly higher plasma α-tocopherol than low all-rac-α-tocopherol supplement (<2.5 mg/d) users (34 vs. 25 μmol/L, P = 0.01). ROC analysis demonstrated an ability to distinguish between RRR- and all-rac-α-tocopherol consumers, which may be useful in investigating the potential effect of RRR- and all-rac-α-tocopherol intake on health., Conclusions: This study demonstrated that the percentage of the population meeting the vitamin E recommendation was unaffected when all-rac-α-tocopherol intake was corrected for α-tocopherol equivalent bioactivity. all-rac-α-Tocopherol intake led to a decrease in the percentage of plasma RRR-α-tocopherol relative to RRR-α-tocopherol intake., (© 2015 American Society for Nutrition.)

Background: Meal pattern analysis can be complex because of the large variability in meal consumption. The use of aggregated, generic meal data may address some of these issues., Objective: The objective was to develop a meal coding system and use it to explore meal patterns., Design: Dietary data were used from the National Adult Nutrition Survey (2008-2010), which collected 4-d food diary information from 1500 healthy adults. Self-recorded meal types were listed for each food item. Common food group combinations were identified to generate a number of generic meals for each meal type: breakfast, light meals, main meals, snacks, and beverages. Mean nutritional compositions of the generic meals were determined and substituted into the data set to produce a generic meal data set. Statistical comparisons were performed against the original National Adult Nutrition Survey data. Principal component analysis was carried out by using these generic meals to identify meal patterns., Results: A total of 21,948 individual meals were reduced to 63 generic meals. Good agreement was seen for nutritional comparisons (original compared with generic data sets mean ± SD), such as fat (75.7 ± 29.4 and 71.7 ± 12.9 g, respectively, P = 0.243) and protein (83.3 ± 26.9 and 80.1 ± 13.4 g, respectively, P = 0.525). Similarly, Bland-Altman plots demonstrated good agreement (<5% outside limits of agreement) for many nutrients, including protein, saturated fat, and polyunsaturated fat. Twelve meal types were identified from the principal component analysis ranging in meal-type inclusion/exclusion, varying in energy-dense meals, and differing in the constituents of the meals., Conclusions: A novel meal coding system was developed; dietary intake data were recoded by using generic meal consumption data. Analysis revealed that the generic meal coding system may be appropriate when examining nutrient intakes in the population. Furthermore, such a coding system was shown to be suitable for use in determining meal-based dietary patterns., (© 2015 American Society for Nutrition.)

Background: Ireland has traditionally operated a liberal policy of voluntary fortification, but little is known about how this practice, along with supplement use, affects population intakes and status of folate and vitamin B-12., Objective: The aim was to examine the relative impact of voluntary fortification and supplement use on dietary intakes and biomarker status of folate and vitamin B-12 in Irish adults., Design: Folic acid and vitamin B-12 from fortified foods and supplements were estimated by using brand information for participants from the cross-sectional National Adult Nutrition Survey 2008-2010. Dietary and biomarker values were compared between 6 mutually exclusive consumption groups formed on the basis of folic acid intake., Results: The consumption of folic acid through fortified foods at low, medium, and high levels of exposure [median (IQR) intakes of 22 (13, 32), 69 (56, 84), and 180 (137, 248) μg/d, respectively]; from supplements [203 (150, 400) μg/d]; or from both sources [287 (220, 438) μg/d] was associated with significantly higher folate intakes and status compared with nonconsumption of folic acid (18% of the population). Median (IQR) red blood cell (RBC) folate increased significantly from 699 (538, 934) nmol/L in nonconsumers to 1040 (83, 1390) nmol/L in consumers with a high intake of fortified foods (P < 0.001), with further nonsignificant increases in supplement users. Supplement use but not fortification was associated with significantly higher serum vitamin B-12 concentrations relative to nonconsumers (P < 0.001). Two-thirds of young women had suboptimal RBC folate for protection against neural tube defects (NTDs); among nonconsumers of folic acid, only 16% attained optimal RBC folate., Conclusions: The consumption of voluntarily fortified foods and/or supplement use was associated with significantly higher dietary intakes and biomarker status of folate in Irish adults. Of concern, the majority of young women remain suboptimally protected against NTDs., (© 2015 American Society for Nutrition.)

Background: The association between sugar-sweetened beverages (SSBs) and health risks remains controversial. To clarify proposed links, reliable and accurate dietary assessment methods of food intakes are essential., Objective: The aim of this present work was to use a metabolomics approach to identify a panel of urinary biomarkers indicative of SSB consumption from a national food consumption survey and subsequently validate this panel in an acute intervention study., Design: Heat map analysis was performed to identify correlations between ¹H nuclear magnetic resonance (NMR) spectral regions and SSB intakes in participants of the National Adult Nutrition Survey (n = 565). Metabolites were identified and receiver operating characteristic (ROC) analysis was performed to assess sensitivity and specificity of biomarkers. The panel of biomarkers was validated in an acute study (n = 10). A fasting first-void urine sample and postprandial samples (2, 4, 6 h) were collected after SSB consumption. After NMR spectroscopic profiling of the urine samples, multivariate data analysis was applied., Results: A panel of 4 biomarkers-formate, citrulline, taurine, and isocitrate-were identified as markers of SSB intake. This panel of biomarkers had an area under the curve of 0.8 for ROC analysis and a sensitivity and specificity of 0.7 and 0.8, respectively. All 4 biomarkers were identified in the SSB sample. After acute consumption of an SSB drink, all 4 metabolites increased in the urine., Conclusions: The present metabolomics-based strategy proved to be successful in the identification of SSB biomarkers. Future work will ascertain how to translate this panel of markers for use in nutrition epidemiology., (© 2015 American Society for Nutrition.)

Reliable dietary assessments are essential when attempting to understand the complex links between diet and health. Traditional methods for collecting dietary exposure can be unreliable, therefore there is an increasing interest in identifying biomarkers to provide a more accurate measurement. Metabolomics is a technology that offers great promise in this area. The aim of this study was to use a multivariate statistical strategy to link lipidomic patterns with dietary data in an attempt to identify dietary biomarkers. We assessed the relationship between lipidomic profiles and dietary data in volunteers (n=34) from the Metabolic Challenge Study (MECHE). Principal component analysis (PCA), linear regression and receiver operating characteristic (ROC) analysis were used to (1) reduce the lipidomic data into lipid patterns (LPs), (2) investigate relationships between these patterns and dietary data and (3) identify biomarkers of dietary intake. Our study identified a total of 6 novel LPs. LP1 was highly predictive of dietary fat intake (area under the curve AUC=0.82). A random forest (RF) classification model used to discriminate between low and high consumers resulted with an error rate of >10%, with a panel of six metabolites identified as the most predictive. LP4 was highly predictive of alcohol intake (AUC=0.81) with lysophosphatidylcholine alkyl C18:0 (LPCeC18:0) identified as a potential biomarker of alcohol consumption. LP6 had a reasonably good ability to predict dietary fish intake (AUC=0.76), with lysophosphatidylethanolamine acyl C18:2 (LPEaC18:2) phoshatidylethanolamine diaclyl C38:4 (PEaaC38:4) identified as potential biomarkers. The identification of these LPs and specific biomarkers will help in better classifying a persons dietary intake and in turn will improve the assessment of the relationship between diet and disease. Linking these LPs and specific biomarkers with health parameters will be an important future step., (Copyright © 2014 Elsevier B.V. All rights reserved.)

In recent years, there has been a great expansion in the nature of new technologies for the study of all biologic subjects at the molecular and genomic level and these have been applied to the field of human nutrition. The latter has traditionally relied on a mix of epidemiologic studies to generate hypotheses, dietary intervention studies to test these hypotheses, and a variety of experimental approaches to understand the underlying explanatory mechanisms. Both the novel and traditional approaches have begun to carve out separate identities vís-a-vís their own journals, their own international societies, and their own national and international symposia. The present review draws on the advent of large national nutritional phenotype databases and related technological developments to argue the case that there needs to be far more integration of molecular and public health nutrition. This is required to address new joint approaches to such areas as the measurement of food intake, biomarker discovery, and the genetic determinants of nutrient-sensitive genotypes and other areas such as personalized nutrition and the use of new technologies with mass application, such as in dried blood spots to replace venipuncture or portable electronic devices to monitor food intake and phenotype. Future development requires the full integration of these 2 disciplines, which will provide a challenge to both funding agencies and to university training of nutritionists., (© 2014 American Society for Nutrition.)

Background: The Vitamin D Standardization Program (VDSP) has developed protocols for standardizing procedures of 25-hydroxyvitamin D [25(OH)D] measurement in National Health/Nutrition Surveys to promote 25(OH)D measurements that are accurate and comparable over time, location, and laboratory procedure to improve public health practice., Objective: We applied VDSP protocols to existing ELISA-derived serum 25(OH)D data from the Irish National Adult Nutrition Survey (NANS) as a case-study survey and evaluated their effectiveness by comparison of the protocol-projected estimates with those from a reanalysis of survey serums by using liquid chromatography-tandem mass spectrometry (LC-tandem MS)., Design: The VDSP reference system and protocols were applied to ELISA-based serum 25(OH)D data from the representative NANS sample (n = 1118). A reanalysis of 99 stored serums by using standardized LC-tandem MS and resulting regression equations yielded predicted standardized serum 25(OH)D values, which were then compared with LC-tandem MS reanalyzed values for all serums., Results: Year-round prevalence rates for serum 25(OH)D concentrations <30, <40, and <50 nmol/L were 6.5%, 21.9%, and 40.0%, respectively, via original ELISA measurements and 11.4%, 25.3%, and 43.7%, respectively, when VDSP protocols were applied. Differences in estimates at <30- and <40-nmol/L thresholds, but not at the <50-nmol/L threshold, were significant (P < 0.05). A reanalysis of all serums by using LC-tandem MS confirmed prevalence estimates as 11.2%, 27.2%, and 45.0%, respectively. Prevalences of serum 25(OH)D concentrations >125 nmol/L were 1.2%, 0.3%, and 0.6% by means of ELISA, VDSP protocols, and LC-tandem MS, respectively., Conclusion: VDSP protocols hold a major potential for national nutrition and health surveys in terms of the standardization of serum 25(OH)D data.

Background: The response to dietary fat plays a key role in metabolic health. Although this can vary widely between individuals, variation within an individual and the associated contribution of phenotypic and genotypic factors to this variation are less defined., Objectives: The objectives were to quantify within-person variation in triacylglycerol response by means of a novel variation score (S(v)) and to explore the phenotypic and genotypic factors associated with this score., Design: Two consecutive 5-h oral-lipid-tolerance tests (OLTTs) were conducted in 51 healthy adults aged 18-60 y with a BMI (in kg/m²) of 18.5 to 49.8. Detailed body composition, physical function, biochemistry, and genotype data were gathered., Results: The postprandial triacylglycerol response profile did not differ (P = 0.64) across OLTTs for the group; nor did average concentrations of functional markers apolipoprotein C2 (P = 0.73) and apolipoprotein C3 (P = 0.74). S(v) was low in most (82%) of the adults and was significantly (P < 0.05) associated with age, fasting triacylglycerol, triacylglycerol AUC, and fasting nonessential fatty acids. Significant associations were also observed between S(v) and single nucleotide polymorphisms in 7 genes (APOA1, IL1α, IL1β, TLR4, TCF7L2, CCK1Rec, and STAT3) after correction for phenotypic differences., Conclusions: This work showed that the within-person variability in postprandial lipemic response is low in most healthy adults. It also showed that variability in this response is associated with a defined set of phenotypic and genotypic characteristics.

Background: It has been suggested that metabolomics could play a role in dietary assessment and identification of novel biomarkers of dietary intake., Objective: This study examined the link between habitual dietary patterns and metabolomic profiles., Design: A total of 160 volunteers participated in a double-blind, randomized, placebo-controlled dietary intervention. We collected biofluids and recorded 3-d food diaries. Food data were reduced to 33 food groups, and a k-means cluster analysis was performed to identify dietary patterns. (1)H Nuclear magnetic resonance (NMR) spectra were acquired for plasma and urine samples, and gas chromatography was used for plasma fatty acid profiling., Results: Cluster analysis identified 3 distinct dietary patterns on the basis of the energy contribution of different food groups. Dietary clusters were reflected in plasma fatty acid profiles and in metabolomic data. (1)H NMR spectra of urine allowed the identification of metabolites associated with different dietary patterns. Several of the metabolites identified were linked to the intake of specific food groups; in particular, there was a positive association between O-acetylcarnitine and phenylacetylglutamine and red-meat and vegetable intakes, respectively., Conclusions: Habitual dietary patterns are shown in metabolomic data. This approach successfully identified potential biomarkers of red-meat and vegetable intakes.

Dietary fat intake plays a critical role in the development of metabolic syndrome (MetS). This study addressed the hypothesis that dietary fat quantity and quality may differentially modulate postprandial lipoprotein metabolism in MetS patients. A multi-center, parallel, randomized, controlled trial conducted within the LIPGENE study randomly assigned MetS patients to 1 of 4 diets: high-SFA [HSFA; 38% energy (E) from fat, 16% E as SFA], high-monounsaturated fatty acid [HMUFA; 38% E from fat, 20% E as MUFA], and 2 low-fat, high-complex carbohydrate [LFHCC; 28% E from fat] diets supplemented with 1.24 g/d of long-chain (LC) (n-3) PUFA (ratio 1.4 eicosapentaenoic acid:1 docosahexaenoic acid) or placebo (1.24 g/d of high-oleic sunflower-seed oil) for 12 wk each. A fat challenge with the same fat composition as the diets was conducted pre- and postintervention. Postprandial total cholesterol, triglycerides (TG), apolipoprotein (apo) B, apo B-48, apo A-I, LDL-cholesterol, HDL-cholesterol and cholesterol, TG, retinyl palmitate, and apo B in TG-rich lipoproteins (TRL; large and small) were determined pre- and postintervention. Postintervention, postprandial TG (P < 0.001) and large TRL-TG (P = 0.009) clearance began earlier and was faster in the HMUFA group compared with the HSFA and LFHCC groups. The LFHCC (n-3) group had a lower postprandial TG concentration (P < 0.001) than the other diet groups. Consuming the LFHCC diet increased the TG (P = 0.04), large TRL-TG (P = 0.01), TRL-cholesterol (P < 0.001), TRL-retinyl palmitate (P = 0.001), and TRL-apo B (P = 0.002) area under the curve compared with preintervention values. In contrast, long-term ingestion of the LFHCC (n-3) diet did not augment postprandial TG and TRL metabolism. In conclusion, postprandial abnormalities associated with MetS can be attenuated with LFHCC (n-3) and HMUFA diets. The adverse postprandial TG-raising effects of long-term LFHCC diets may be avoided by concomitant LC (n-3) PUFA supplementation to weight-stable MetS patients.

Long-chain acyl CoA synthetase 1 (ACSL1) plays an important role in fatty acid metabolism and triacylglycerol (TAG) synthesis. Disturbance of these pathways may result in dyslipidemia and insulin resistance, hallmarks of the metabolic syndrome (MetS). Dietary fat is a key environmental factor that may interact with genetic determinants of lipid metabolism to affect MetS risk. We investigated the relationship between ACSL1 polymorphisms (rs4862417, rs6552828, rs13120078, rs9997745, and rs12503643) and MetS risk and determined potential interactions with dietary fat in the LIPGENE-SU.VI.MAX study of MetS cases and matched controls (n = 1,754). GG homozygotes for rs9997745 had increased MetS risk {odds ratio (OR) 1.90 [confidence interval (CI) 1.15, 3.13]; P = 0.01}, displayed elevated fasting glucose (P = 0.001) and insulin concentrations (P = 0.002) and increased insulin resistance (P = 0.03) relative to the A allele carriers. MetS risk was modulated by dietary fat, whereby the risk conferred by GG homozygosity was abolished among individuals consuming either a low-fat (<35% energy) or a high-PUFA diet (>5.5% energy). In conclusion, ACSL1 rs9997745 influences MetS risk, most likely via disturbances in fatty acid metabolism, which was modulated by dietary fat consumption, particularly PUFA intake, suggesting novel gene-nutrient interactions.

The leptin receptor (LEPR) is associated with insulin resistance, a key feature of metabolic syndrome (MetS). Gene-fatty acid interactions may affect MetS risk. The objective was to investigate the relationship among LEPR polymorphisms, insulin resistance, and MetS risk and whether plasma fatty acids, a biomarker of dietary fatty acids, modulate this. LEPR polymorphisms (rs10493380, rs1137100, rs1137101, rs12067936, rs1805096, rs2025805, rs3790419, rs3790433, rs6673324, and rs8179183), biochemical measurements, and plasma fatty acid profiles were determined in the LIPGENE-SU.VI.MAX study of MetS cases and matched controls (n = 1754). LEPR rs3790433 GG homozygotes had increased MetS risk compared with the minor A allele carriers [odds ratio (OR) = 1.65; 95% CI: 1.05-2.57; P = 0.028], which may be accounted for by their increased risk of elevated insulin concentrations (OR 2.40; 95% CI: 1.28-4.50; P = 0.006) and insulin resistance (OR = 2.15; 95% CI: 1.18-3.90; P = 0.012). Low (less than median) plasma (n-3) and high (n-6) PUFA status exacerbated the genetic risk conferred by GG homozygosity to hyperinsulinemia (OR 2.92-2.94) and insulin resistance (OR 3.40-3.47). Interestingly, these associations were abolished against a high (n-3) or low (n-6) PUFA background. Importantly, we replicated some of these findings in an independent cohort. Homozygosity for the LEPR rs3790433 G allele was associated with insulin resistance, which may predispose to increased MetS risk. Novel gene-nutrient interactions between LEPR rs3790433 and PUFA suggest that these genetic influences were more evident in individuals with low plasma (n-3) or high plasma (n-6) PUFA.

Background: At present, the analysis of dietary patterns is based on the intake of individual foods. This article demonstrates how a coding system at the meal level might be analyzed by using data mining techniques., Objective: The objective was to evaluate the usability of supervised data mining methods to predict an aspect of dietary quality based on dietary intake with a food-based coding system and a novel meal-based coding system., Design: Food consumption databases from the North-South Ireland Food Consumption Survey 1997-1999 were used. This was a randomized cross-sectional study of 7-d recorded food and nutrient intakes of a representative sample of 1379 Irish adults. Meal definitions were recorded by the respondent. A healthy eating index (HEI) score was developed. Artificial neural networks (ANNs) and decision trees were used to predict quintiles of the HEI based on combinations of foods consumed at breakfast and main meals., Results: This study applied both data mining techniques to the food and meal-based coding systems. The ANN had a slightly higher accuracy than did the decision tree in relation to its ability to predict HEI quintiles 1 and 5 based on the food coding system (78.7% compared with 76.9% and 71.9% compared with 70.1%, respectively). However, the decision tree had higher accuracies than did the ANN on the basis of the meal coding system (67.5% compared with 54.6% and 75.1% compared with 72.4%, respectively)., Conclusions: ANNs and decision trees were successfully used to predict an aspect of dietary quality. However, further exploration of the use of ANNs and decision trees in dietary pattern analysis is warranted.

Background: Diversity in dietary intake contributes to variation in human metabolomic profiles and artifacts from acute dietary intake can affect metabolomics data., Objective: We investigated the role of dietary phytochemicals on shaping human urinary metabolomic profiles., Design: First void urine samples were collected from 21 healthy volunteers (12 women, 9 men) following their normal diet (ND), a 2-d low-phytochemical diet (LPD), or a 2-d standard phytochemical diet (SPD). Nutrient intake was assessed during the study. Urine samples were analyzed by using (1)H nuclear magnetic resonance spectroscopy ((1)H NMR) and mass spectrometry (MS), which was followed by multivariate data analysis., Results: Macronutrient intake did not change throughout the study. Partial least-squares-discriminant analysis indicated a clear distinction between the LPD samples and the ND and SPD samples, relating to creatinine and methylhistidine excretion after the LPD and hippurate excretion after the ND and SPD. The predictive power of the LPD versus the ND model was 74 +/- 3% and 82 +/- 6% with the (1)H NMR and MS data sets, respectively. The predictive power of the LPD versus the SPD model was 83 +/- 8% and 69 +/- 4% for the (1)H NMR and MS data sets respectively. A cross platform comparison of both data sets by co-inertia analysis showed a similar distinction between the LPD and SPD., Conclusions: Acute changes in urinary metabolomic profiles occur after the consumption of dietary phytochemicals. Dietary restrictions in the 24 h before sample collection may reduce diversity in phytochemical intakes and therefore reduce variation and improve data interpretation in metabolomics studies using urine.

Background: Metabolomics in human nutrition research is faced with the challenge that changes in metabolic profiles resulting from diet may be difficult to differentiate from normal physiologic variation., Objective: We assessed the extent of intra- and interindividual variation in normal human metabolic profiles and investigated the effect of standardizing diet on reducing variation., Design: Urine, plasma, and saliva were collected from 30 healthy volunteers (23 females, 7 males) on 4 separate mornings. For visits 1 and 2, free food choice was permitted on the day before biofluid collection. Food choice on the day before visit 3 was intended to mimic that for visit 2, and all foods were standardized on the day before visit 4. Samples were analyzed by using 1H nuclear magnetic resonance spectroscopy followed by multivariate data analysis., Results: Intra- and interindividual variations were considerable for each biofluid. Visual inspection of the principal components analysis scores plots indicated a reduction in interindividual variation in urine, but not in plasma or saliva, after the standard diet. Partial least-squares discriminant analysis indicated time-dependent changes in urinary and salivary samples, mainly resulting from creatinine in urine and acetate in saliva. The predictive power of each model to classify the samples as either night or morning was 85% for urine and 75% for saliva., Conclusions: Urine represented a sensitive metabolic profile that reflected acute dietary intake, whereas plasma and saliva did not. Future metabolomics studies should consider recent dietary intake and time of sample collection as a means of reducing normal physiologic variation.

Metabolomics has been widely adopted in pharmacology and toxicology but is relatively new in human nutrition. The ultimate goal, to understand the effects of exogenous compounds on human metabolic regulation, is similar in all 3 fields. However, the application of metabolomics to nutritional research will be met with unique challenges. Little is known of the extent to which changes in the nutrient content of the human diet elicit changes in metabolic profiles. Moreover, the metabolomic signal from nutrients absorbed from the diet must compete with the myriad of nonnutrient signals that are absorbed, metabolized, and secreted in both urine and saliva. The large-bowel microflora also produces significant metabolic signals that can contribute to and alter the metabolome of biofluids in human nutrition. Notwithstanding these possible confounding effects, every reason exists to be optimistic about the potential of metabolomics for the assessment of various biofluids in nutrition research. This potential lies both in metabolic profiling through the use of pattern-recognition statistics on assigned and unassigned metabolite signals and in the collection of comprehensive data sets of identified metabolites; both objectives have the potential to distinguish between different dietary treatments, which would not have been targeted with conventional techniques. The latter objective sets out a well-recognized challenge to modern biology: the development of libraries of small molecules to aid in metabolite identification. The purpose of the present review was to highlight some early challenges that need to be addressed if metabolomics is to realize its great potential in human nutrition.

Macrophage cholesterol homeostasis is a key process involved in the initiation and progression of atherosclerosis. Peroxisome proliferator-activated receptors (PPARs) regulate the transcription of the genes involved in cholesterol homeostasis and thus represent an important therapeutic target in terms of reducing atherosclerosis. Conjugated linoleic acid (CLA) is a potent anti-atherogenic dietary fatty acid in animal models of atherosclerosis and is capable of activating PPARs in vitro and in vivo. Therefore, this study examined whether the anti-atherogenic effects of CLA in vivo could be ascribed to altered cholesterol homeostasis in macrophages and macrophage derived foam cells. Of several genes that regulate cholesterol homeostasis investigated, CLA had most effect on the class B scavenger receptor CD36. The cis-9,trans-11 CLA (c9,t11-CLA) and trans-10,cis-12 CLA (t10,c12-CLA) isomers augmented CD36 mRNA expression (P<0.001). Confocal laser microscopy characterised the three-dimensional expression patterns of CD36 in THP-1 macrophages. Linoleic acid, CLA and the PPARgamma ligand rosiglitazone increased discrete cell surface CD36 localisation, with a heterogeneous punctate pattern of expression. In agreement with the observed increases in CD36 mRNA and cell surface expression, intracellular cholesterol concentrations were greater in macrophages exposed to linoleic acid and CLA. Further analysis of cholesterol metabolism showed that CLA had no effect on THP-1 derived foam cell cholesterol efflux to apo AI. Thus, altered cholesterol homeostasis in the macrophage may not explain the anti-atherogenic effects of CLA observed in vivo.

Although the amino acid composition of almost all food proteins is known, estimating the amino acid intake from the diet is extremely difficult because of the lack of available data. A conservative approach would be to determine the population distribution of protein intake, select the 97.5(th) or higher percentile of intake, assume all comes from the target protein, and estimate exposure to some specific amino acid. Any number of dietary survey methodologies could be used to conduct such a conservative approach. However, given the great variety of brands of food supplements, estimates of amino acid intakes from supplements are problematic. Firstly, few studies include supplements in their target nutrient sources because brand-level data would need to be retained and nutritional composition data would need to be recorded. Probabilistic modeling offers some solution provided some basic data are gathered. The percentage of the population regularly taking supplements and the frequency of consumption must be known. Therefore, data on the dietary supplement market would need to be known including the percent of brands containing amino acids and if possible specific amino acids together with concentrations. A probabilistic model as follows would ensue: probability of being a consumer of amino acid supplements; probability distribution function of frequency of use of supplements; probability distribution function of dose per eating occasion; market characteristics; probability distribution function for dietary amino acid intake. Using multiple iterations and perhaps bootstrapping on some elements of the model, fully worst-case model scenarios of exposure could be computed.

Conjugated linoleic acid (CLA) has profound effects on hepatic and adipocyte lipid metabolism, but little is known about its effects on intestinal lipid metabolism. We investigated the acute (22 h) and acute-after-chronic (22 h after 19 d) effects of trans-10, cis-12 CLA (t10,c12-CLA) and cis-9, trans-11 CLA (c9, t11-CLA) on triacylglycerol (TAG)-rich lipoprotein (TRL) metabolism, de novo TAG, phospholipid (PL) ((14)C-glycerol) and apolipoprotein B (apoB) ((35)S-methionine) synthesis and secretion, in the colon carcinoma (Caco-2) cell model of intestinal lipoprotein metabolism. Acute treatment with either CLA isomer did not affect TRL metabolism. However, chronic t10,c12-CLA and c9,t11-CLA supplementation followed by acute palmitic acid (PA) treatment increased the ratio of cellular to secreted de novo TAG (cTAG/sTAG) (P < or = 0.03) as a result of increased cellular de novo TAG levels. Chronic Caco-2 cell t10,c12-CLA supplementation, prior to the acute oleic acid (OA) treatment, significantly increased (P = 0.005) the ratio of cellular de novo TAG to de novo PL (cTAG/cPL), to a greater extent than that following chronic linoleic acid (LA) (P = 0.001) or c9,t11-CLA supplementation (P = 0.005). Again, this effect was attributed to increased cellular de novo TAG synthesis. Neither CLA isomer affected the ratio of secreted de novo TAG to de novo PL (sTAG/sPL). No effects on Caco-2 cell apoB synthesis and secretion were observed after acute or chronic CLA treatments. In conclusion, chronic t10,c12-CLA supplementation modulated intestinal TRL metabolism, by increasing cellular de novo TAG synthesis but had no effect on de novo TAG secretion in Caco-2 cells.

An observational, cross-sectional study was conducted in a representative sample of the European Union (7155 men and 8077 women) to calculate the underestimation of body weight as assessed by body image among the overweight and obese population and identify the associated factors to this behavior. Participants were older than 15 years and they were living in the 15 European Union countries. Body mass index (BMI) was grouped into 4 categories using the cutpoints established by the WHO, while perceived body image (PBI) was assessed using a nine-silhouettes drawing. The degree of underestimation between PBI as compared to BMI was identified in overweight and obesity categories of BMI. A multivariable logistic regression model for each gender was used to adjust for potentially confounding variables. Men classified themselves worse than women, being more likely to underestimate their body weight (65.2% of men underestimated their weight vs 32.2% women), regardless of other socioeconomic and attitudinal variables. The greatest degree of underestimation was observed in Mediterranean individuals (68.7% of men and 37.9% of women underestimated their weight). The subjects in the 'maintenance' stage of physical activity tended more often to wrongly select their actual image (71.8% for men and 38.7% for women).

Elevated plasma triacylglycerol concentrations have been associated with increased risk of coronary heart disease (CHD). In the past, the epidemiologic evidence about the causal role of triacylglycerols in CHD has not been well regarded, but recent prospective evidence shows that nonfasting plasma triacylglycerol concentration is a strong and independent predictor of future myocardial infarction. Elevated plasma triacylglycerol concentrations are associated with other CHD risk factors, namely reduced HDL-cholesterol concentrations and a preponderance of highly atherogenic, small, dense LDL particles. Plasma triacylglycerol concentrations increase after the ingestion of a fat-containing meal, and elevated postprandial triacylglycerolemia leads to a series of metabolic reactions that reduce HDL-cholesterol concentrations and promote the formation of small, dense LDL particles. The magnitude of the postprandial response is largely determined by fasting plasma triacylglycerol concentrations. Metabolism of plasma triacylglycerols also influences postprandial factor VII activation and the postprandial lipemic responsiveness to dietary cholesterol. Therefore, dietary factors that improve fasting plasma triacylglycerol concentrations must have a role in a healthy diet. Eicosapentaenoic and docosahexaenoic acids are n-3 polyunsaturated fatty acids (PUFAs) in fish oil that effectively reduce plasma triacylglycerol concentrations. Because n-3 PUFAs are effective at low doses (1 g n-3 PUFA/d), they provide a realistic option for the optimization of plasma triacylglycerol metabolism.

Background: This study was an extension of a previous study that showed different lipemic responses to standard test meals in subjects from southern and northern Europe., Objective: The aim was to determine in 32 healthy young men from northern and southern Europe whether differences in the secretion of insulin and glucose-dependent insulinotrophic polypeptide (GIP) might explain these findings through the actions of these hormones on lipoprotein lipase., Design: We investigated in a randomized, single-blind, crossover study the effects of 2 test meals of identical macronutrient composition but different saturated fatty acid (SFA) and monounsaturated fatty acid (MUFA) contents on postprandial GIP, insulin, the ratio of incremental triacylglycerol to apolipoprotein B-48 (a marker of chylomicron size), and the activity of postheparin lipases., Results: Fasting and postprandial GIP concentrations and postheparin hepatic lipase activities were significantly higher in the southern Europeans (P < 0.001 and P < 0.02, respectively). Lipoprotein lipase activity after the SFA-rich meal was significantly higher in the northern Europeans (P < 0.01). HL activity 9 h after the SFA-rich meal and the area under the curve (AUC) for the postprandial insulin response correlated with the AUC for the postprandial GIP response [r = 0.44 (P < 0.04) and r = 0.46 (P < 0.05), respectively]. There were no significant differences in chylomicron size between the 2 groups for either meal, but when the groups were combined there was a significant difference in chylomicron size between the SFA- and MUFA-rich meals (P < 0.05), which could be due to the formation of larger chylomicrons after the MUFA-rich meal., Conclusion: The significantly higher GIP and insulin responses and HL activities in southern Europeans may provide an explanation for our previous report of attenuated postprandial triacylglycerol and apolipoprotein B-48 responses in them.

Given the wide array of demands on food intake databases for estimating food chemical intake, there is no possibility of an all-embracing methodology. Whereas initial estimates should always begin at a crude level, when more refined estimates are needed methodologies range from indirect (household, national level) to direct (retrospective or prospective, individual level). The indirect methods cannot provide data on consumer-only intakes but percentage households purchasing can be used as a surrogate for percentage individuals consuming. For the direct methods, the greater the duration, the higher the estimate of percentage consumers and the lower the estimate of intakes among consumers only. In considering excursions above the acceptable daily intake (ADI), databases need to be constructed which allow estimates of intake per eating occasion, where intakes at each eating occasion through the day or days are collapsed into intake per day or week, and the possibility of measuring excursions above the ADI is lost.

Although the beneficial effects of Mediterranean-type diets, which are rich in olive oil, a good source of monounsaturated fatty acids (MUFAs), are generally accepted, little is known about the effects of long-term dietary MUFA intake on postprandial lipoprotein metabolism and hemostasis. This study used a single-blind, randomized, crossover design to investigate the relative effects of a long-term dietary olive oil intervention and a control [saturated fatty acid (SFA)-enriched] diet on postprandial triacylglycerol metabolism and factor VII activity. The postprandial response to a standard test meal was investigated in 23 healthy men who adhered to both diets for 8 wk. cis-MUFAs were successfully substituted for SFAs in the MUFA diet without affecting total dietary fat or energy intakes. The long-term dietary MUFA intervention significantly reduced plasma and LDL-cholesterol concentrations (P = 0.01). Postprandial triacylglycerol concentrations were significantly greater in the early postprandial period after the MUFA diet (P = 0.003). Postprandial factor VII activation and the concentration of the factor VII antigen were significantly lower after the MUFA diet (P = 0.04 and P = 0.006, respectively). This study showed that isoenergetic substitution of MUFAs for SFAs reduces plasma cholesterol and reduces the degree of postprandial factor VII activation. The alterations in the postprandial triacylglycerol response suggest a greater rate of dietary fat absorption and postprandial triacylglycerol metabolism after a diet rich in MUFAs. This study presents new insights into the biochemical basis of the beneficial effects associated with long-term dietary MUFA consumption, which may explain the lower rates of coronary mortality in Mediterranean regions.

Postprandial lipaemic responses to two test meals were investigated in 30 Northern (15 British and 15 Irish), and 30 Southern (Greeks from Crete) healthy male Europeans. The meals were a saturated fatty acid (SFA) meal, which resembled the fatty acid composition of an average UK diet, and a monounsaturated fatty acid (MUFA) meal in which the fat consisted of olive oil. Habitual diets of the two groups differed, with higher total fat, (P < 0.03) and MUFA (P < 0.0001) and lower polyunsaturated fatty acid (PUFA) (P < 0.0001) intakes in Southern than Northern Europeans. Levels of total MUFA (P < 0.02) and oleic acid (P < 0.004) were also higher in adipose tissue of Southern in comparison to Northern Europeans. In both European groups there were no significant differences in postprandial triglyceride response between the two meal types, SFA or MUFA. However, Northern and Southern Europeans showed significant differences in their patterns of postprandial response in plasma triglycerides (P < 0.0001), apolipoprotein B-48 (P < 0.0001), NEFA (P < 0.0001), insulin (P < 0.0007), and factor VII activity (P-0.03). In the case of NEFA, areas under the response curve were higher following the SFA than the MUFA meal for both groups, (P < 0.003) and were greater in Southern than Northern Europeans (P < 0.002) and apo B-48 responses were lower (P < 0.005). Some of these differences may reflect differences in fasting levels since fasting apolipoprotein B-48 levels were lower (P < 0.01) and fasting NEFA (P < 0.02) and insulin (P < 0.005) were higher in the Southern than in the Northern Europeans. In addition, 9 h postprandial post-heparin lipoprotein lipase activity was lower in the Southern than in the Northern Europeans (P < 0.0006). This is the first report of differences in postprandial lipid, factor VII and insulin responses in Southern and Northern Europeans which may be of importance in explaining the different susceptibilities of these two populations to risk of coronary artery disease.

The Maasai of East Africa have been found to have low serum concentrations of cholesterol and a low incidence of cardiovascular disease in spite of apparently very high milk intakes. On that basis it has been frequently suggested that milk contains a "hypocholesterolaemic factor". The hypocholesterolaemia of the Maasai had also been attributed to a genetic adaptation. We feel that the milk intakes reported for the Maasai are excessively high and that the low incidence of cardiovascular diseases and low levels of serum cholesterol may be adequately explained by their variable and generally low energy intakes.

Breeding rabbits were fed diets in which milk protein provided 30% of total energy, both before conception and throughout pregnancy and lactation. Twelve of their offspring were maintained on a similar diet containing 166 g/kg saturated fat. A second group, bred from a colony given a milk protein-free ration, was fed an identical diet. Anti-milk antibodies were higher in the group reared from mothers not exposed to dietary milk. There were no differences in circulating immune complexes and the extent of aortic atherosclerosis was very similar after one year's feeding. It was concluded that the magnitude of the immune response to dietary milk protein was insufficient to influence the development of experimental atherosclerosis.

Coronary arteriography was used to identify 37 patients with triple coronary vessel disease and 37 patients with no evidence of occlusive coronary atheroma. Two techniques were used to analyse serum levels of antibodies to heated milk protein, passive haemagglutination and enzyme-linked immunosorbent assay. No significant differences in these antibodies were observed between triple and zero vessel disease patients. Even after applying ECG evidence of myocardial infarction as a further criterion for classifying patients, no evidence was obtained to support the hypothesis that serum antibodies to heated milk protein are elevated in coronary heart disease.

Four groups of 6 New Zealand white rabbits were fed for 60 days on a commercial rabbit diet supplemented (60 g/kg) with 1 of 4 sources of fat: corn oil, linseed oil and fish oil, which respectively provided rich sources of linoleic acid (C18:2, n-6), alpha-linolenic acid (C18:3, n-3) and eicosapentaenoic acid (C20:5, n-3), and coconut oil, low in all polyunsaturated fatty acids (PUFA). Platelet-rich plasma was prepared and aggregation induced by ADP (final concentration 0.29-74.3 microM), collagen (2.5-20 micrograms/400 microliters, final concentration) and 2.5 U bovine thrombin, and recorded with a Peyton aggregometer. Platelet aggregation induced by both thrombin and collagen was significantly lower with either n-3 PUFA (fish or linseed oil) than with corn oil (n-6 PUFA) or the low PUFA coconut oil. ADP-induced platelet aggregation was significantly reduced only in animals fed fish-oil. Changes in platelet aggregation were accompanied by increased platelet lipid content of C20:5, n-3 and decreased content of C20:4, n-6, with little change in platelet total C20 fatty acids. Platelet levels of C20:5, n-3 were significantly increased with both the preformed C20:5, n-3 and its precursor C18:3, n-3 in the diet. However, aortic lipid accumulation of C20:5 only occurred with rabbits fed on fish oil. It was concluded that, for collagen and thrombin induced aggregation, C18:3, n-3 and C20:5, n-3 were equally antiaggregatory in rabbits. The implications of this in community nutrition programmes are discussed.

The effect of varying the compositions of dietary proteins on the relative cholesterolaemic effects of animal and vegetable proteins was investigated in rabbits. In experiments using high fat diets, the amino acid compositions of dietary proteins (soya or casein) were altered by blending them 1:1 (w/w) with gelatin. This reduced the differences in amino acid compositions and also made soya more and casein less hypercholesterolaemic. In experiment 2a, soya protein was compared with dried skim milk in low fat diets and in experiment 2b, these proteins were supplemented with lysine or arginine, respectively, so that the lysine:arginine ratio of soya was similar to dried skim milk and vice-versa. Serum cholesterol was significantly higher in milk-fed than soya-fed rabbits and was not influenced by reversing the lysine:arginine ratio. In the three experiments, parameters of cholesterol kinetics were estimated from the die-away curve of injected [4 14-C]cholesterol. There were no significant effects of diet on the parameters of cholesterol kinetics. It was concluded that the lysine:arginine ratio of the diet is not the major determinant of the cholesterolaemic properties of proteins, but that the overall amino acid composition is primarily concerned.

The effect of dietary fish oil (rich in n-3 polyunsaturated fatty acids (PUFA], corn oil (rich in n-6 PUFA) and coconut oil (low in n-3 and n-6 PUFA) on the induction of atherosclerosis by serum sickness in rabbits was investigated over a 12-month period. Dietary fish oil led to a significant increase in the level of eicosapentaenoic acid (EPA) in all platelet phospholipid fractions and to a significant reduction in the level of platelet phosphatidylethanolamine arachidonic acid (AA). In aortic total phospholipids, rabbits given fish oil showed a significant reduction in AA and a significant increase in EPA. Rabbits given fish oil showed significantly lower collagen-induced platelet thromboxane A2 release and aortic production of 6-keto-PGF1 alpha. Serum total immune complex levels and anti-horse serum IgG levels were not influenced by diet. There was a significant reduction in total aortic atherosclerosis in fish oil-fed animals compared with coconut oil fed animals.

Breeding rabbits were fed diets supplemented with 5% milk or soya protein for 1 month before conception and throughout pregnancy and lactation. Four groups of 5 or 6 of their offspring were given high-energy isonitrogenous diets containing either 30% soya or milk protein. Half of the animals from each litter received the same protein as their dams; the remainder were fed the alternative (i.e. dams soya; offspring milk, and vice versa). 0.75% cholesterol was added to the diets between 30 and 120 days after weaning. Animals given the same protein as their dams formed substantially lower amounts of food antigen-specific antibody than rabbits fed a novel protein at weaning but the extent of aortic atherosclerosis was similar in all groups. In a second experiment groups of 5-8 weanling rabbits were fed a cholesterol-free diet containing 30% soya and 17% saturated fat for 1 year. Animals in group 1 were bred from dams given soya but those in groups 2 and 3 were derived from a colony fed a soya-free diet. Rabbits in group 3 were immunised by repeated parenteral injections of soya protein and developed high levels of antisoya antibodies. Group 1 and 2 animals were injected with saline only but antisoya antibodies were substantially higher in animals derived from the soya-free breeding colony. Although serum cholesterol and triglyceride levels were similar in all groups, animals from dams fed soya (group 1) had significantly less aortic atherosclerosis (10% involvement of ascending aorta) than those reared from a colony fed a soya-free diet (group 2; 32%). Parenteral immunization with soya protein (group 3) was not associated with significantly increased atherosclerosis (37%). These findings indicate that perinatal exposure to dietary antigen in rabbits may be important in modulating the systemic immune reaction to food antigens. The magnitude of this systemic response is unlikely to alter the nature or distribution of cholesterol-induced atherosclerosis but may have an important influence on the development of aortic disease produced by prolonged feeding of high-fat, cholesterol-free diets.

Groups of rabbits were fed isonitrogenous diets containing 30% soya, milk or whitefish meal protein and at least 130 g/kg of added fat for 1 year. Mean serum cholesterol values in fish meal animals (13.4 +/- 2.4 mmol/l) were substantially greater than for soya (3.0 +/- 0.2 mmol/l) or milk (4.6 +/- 0.7 mmol/l). Fish meal rabbits developed extensive aortic atherosclerosis (c. 70% surface involvement) which histologically showed both fibrous and foam cell intimal thickening and destruction and calcification of medial elastic tissue. In a second experiment fish meal was given with either saturated (coconut oil) or unsaturated (maize oil) fat. A similar degree of hypercholesterolaemia developed in each group and was rapidly reversed when a soya protein-low fat diet was substituted. This model may therefore be of value in studies of the progression and regression of experimental atherosclerosis.