To investigate the impact of gender and age on metabolic profile of blood plasma of Celiac disease (CeD) patients in comparison to healthy controls using in vitro proton (¹H) nuclear magnetic resonance (NMR) spectroscopy.CeD is more prevalent in women with a female to male ratio between 2:1 and 3:1 and the clinical presentation of CeD is not same in men and women¹. Various physiological factors like gender, age and race influence the metabolism. Therefore, in the present study, the influence of gender and age on metabolic pattern of CeD patients in comparison to controls was investigated using NMR based metabonomics. Thirty treatment-naïve patients with CeD (mean age 25.3±10.9 years; female, n=16 and male, n=14) and 30 healthy controls (mean age 28.8±6.4yrs; female, n=15 and male n=15) were recruited. An informed consent was taken and study was approved by the Institute Ethics Committee. All patients were treated according to standard regimen. Diagnosis of CeD was made on the basis of European Society of Pediatric Gastroenterology Hepatology and Nutrition (ESPGHAN). Blood samples were collected in morning pre-prandial and plasma were separated and stored at -80 °C until NMR analysis. One-dimensional Carr-Purcell-Meiboom-Gill Sequence and two dimensional total correlation spectroscopy experiments were performed at 700 MHz (Agilent, U.S.A.). Student t-test/Wilcoxon rank sum test were used for comparison and a probability <0.05 was considered significant. Partial least squares-discriminant analysis (PLS-DA) was performed using Unscrambler 10.2 (CAMO Software, Oslo, Norway).The concentrations [(μM (mean ±SD)] of leucine, valine and phosphocreatine to creatine ratio were significantly higher in healthy males compared to healthy females (Table 1). Significant differences were seen in concentration of several metabolites between males and females with CeD when patients were categorized in two groups: (a) below, and (b) above the age of 20 yrs (Table 2). The concentrations of acetate, pyruvate, creatine and glycine were significantly higher in males with CeD compared to healthy males (Table 1). While, the levels of β-hydroxybutyrate, glycine and alanine were significantly elevated in females with CeD than healthy females. PLS-DA showed clear distinction between healthy males and healthy females (Fig. 1); males with CeD and healthy males (Fig. 2); and females with CeD and healthy females (Fig. 3). To the best of our knowledge, this the first study that demonstrated the influence of gender and age on the metabolism of CeD patients in comparison to healthy subjects. Significant gender specific and age related changes were seen in healthy subjects as well as in CeD patients. Separate clusters of healthy males and healthy females; males with CeD and healthy males; females with CeD and healthy females were observed on PLS-DA plots. Healthy males had higher concentration of leucine and valine and higher phosphocreatine/creatine ratio compared to healthy females indicating high protein turnover and muscle metabolism in healthy males². Younger males with CeD had higher concentration of acetate, tyrosine, histidine and phenylalanine compared to females with CeD. While among CeD patients above the age of 20 yrs, only glucose and choline were significantly higher in males. Further our data showed that males with CeD had higher concentration of acetate compared to healthy males, while females with CeD showed elevated β-hydroxybutyrate than healthy females. These metabolic differences indicated that carbohydrate metabolism is impaired in CeD patients of both genders, however, fuel preference to meet energy supply was gender specific. Males with CeD used fatty acids while females used ketone bodies as energy substrates. Males with CeD showed higher concentration of pyruvate and creatine and lower phosphocreatine/creatine ratio compared to healthy males, reflecting low energy status. While these metabolites were lower but not significantly different in females with CeD compared to healthy females. Our data also showed higher concentration of gluconeogenic amino acids alanine and glycine in females while only glycine in CeD males compared to healthy females and males, respectively. These findings indicated that these amino acids probably might not have been utilized for gluconeogenesis in CeD. The decreased level of creatinine was observed both in males and females with CeD indicating protein malabsorption. The present study indicated gender-specific fuel preferences and nutritional status in both CeD and healthy subjects, and hence, nutritional recommendations may be based on gender and age for the treatment management of CeD patients.