**Background:** Uzbekistan has high rates of diet-related chronic diseases, potentially linked to high animal fat intake. Sheep meat accounts for approximately one-third of red meat intake in Uzbekistan and is considered health-promoting by locals. Sheep meat contains ~5% fat in muscle, with higher n-3 polyunsaturated fatty acids (2.49±0.99%) and conjugated linoleic acids (10.3±5.2%) compared to beef (1.48±0.02% and 4.9±1.8%, respectively), but also higher total saturated fatty acids (48.3±2.09% vs 43±1.7%). Despite sheep meat's prominence in traditional diets globally, its effect on the human metabolome had not been previously investigated. This study aimed to apply a metabolomics approach to investigate whether sheep meat intake frequency (SMIF) is associated with alterations in fasting blood plasma metabolites and lipoproteins in healthy Uzbek adults.
**Methods:** A total of 263 healthy subjects (149 females, 114 males; age 46.94±15.91 years; BMI 25.83±4.79 kg/m²) were recruited at the Center for Advanced Technologies in Tashkent, Uzbekistan. Exclusion criteria included pregnancy, lactation (up to 6 weeks prior), diagnosed cardiovascular disease, diabetes, cancer, chronic gastrointestinal disorders, use of lipid-lowering/anti-diabetic/anti-hypertensive drugs, or antibiotic treatment within the last three months. Subjects completed a food intake questionnaire including SMIF, total meat intake frequency, and fish intake frequency. Fasting blood plasma samples were collected and analyzed using ¹H NMR spectroscopy (Bruker Avance III 600 MHz). Sixty metabolites and 65 lipoproteins were quantified using SigMa software. Subjects were classified into three groups: high SMIF (>10 times/month, n=80), moderate SMIF (1–10 times/month, n=112), and zero SMIF (<1 time/month, n=71). Statistical analyses included chi-square tests, one-way ANOVA with Benjamini–Hochberg FDR correction (10% FDR, significance threshold p<0.05), multiple linear regression (MLR) adjusted for nationality, sex, BMI, age, total meat intake frequency, and fish intake frequency, as well as PCA, ASCA, and PLS-DA.
**Key Results:** Nationality, sex, BMI, age, total meat intake frequency, and fish intake frequency were significantly confounded with SMIF (p<0.01). The high SMIF group had lower mean blood cholesterol (3.71±0.68 mmol/L) compared to moderate (3.73±0.60 mmol/L) and zero SMIF groups (4.05±0.84 mmol/L; p=0.004). After MLR adjustment for confounders, 12 metabolites remained significantly associated with SMIF. Pyruvic acid (fold change H/Z=0.61, p<0.001, effect 12.62%), phenylalanine (fold change H/Z=0.76, p<0.001, effect 9.04%), ornithine (fold change H/Z=0.68, p<0.001, effect 8.06%), acetic acid (fold change H/Z=0.69, p=0.002, effect 5.92%), formate (fold change H/Z=0.66, p=0.007, effect 4.90%), and proline were significantly lower in the high SMIF group. Choline (fold change H/Z=1.11, p<0.001, effect 8.23%), dimethylglycine/asparagine (fold change H/Z=1.33, p<0.001, effect 9.91%), and serine showed increasing trends with higher SMIF. ASCA revealed SMIF explained 2.8% of total variation in metabolomics data (p=0.0001). PLS-DA achieved a classification error of 17.0% and AUC of 0.86 for distinguishing high vs zero SMIF. For lipoproteins, 14 variables including plasma cholesterol, cholesterol ester, apolipoprotein A1, LDL-4-cholesterol (fold change H/Z=0.76, p<0.001, effect 7.88%), LDL-3-cholesterol ester (fold change H/Z=0.74, p<0.001, effect 7.34%), HDL-cholesterol (fold change H/Z=0.77, p<0.001, effect 6.79%), and HDL-2b-cholesterol (fold change H/Z=0.75, p=0.003, effect 5.09%) were lower in the high SMIF group in MLR, but none remained significant after FDR correction. ASCA showed SMIF explained 4.5% of lipoprotein variation (p<0.0002).
**Clinical Implications:** This first metabolomics investigation of sheep meat intake suggests that frequent consumption is associated with distinct plasma metabolite profiles, particularly lower pyruvate, ornithine, and acetate, and higher choline and dimethylglycine—patterns potentially reflecting differences in energy metabolism, protein turnover, and gut microbiota fermentation. The lower acetate levels in high SMIF subjects may indicate lower dietary fiber intake. The decreasing trend in cholesterol and lipoprotein levels with higher SMIF, though not significant after correction, aligns with previous research suggesting stearic acid in sheep meat may reduce LDL cholesterol. These findings provide a theoretical basis for understanding metabolic changes related to sheep meat consumption but do not establish causality. Limitations include lack of quantified food consumption data, physical activity information, and the observational cross-sectional design. The relatively small sample size and multiple confounding factors likely contributed to non-significant lipoprotein findings after FDR correction. Further longitudinal studies with better dietary characterization are needed to confirm these associations and their clinical relevance for cardiovascular risk.