Human fecal short chain fatty acids determination by SPME-GC-MS: disease monitoring by a chemometric approach in pediatric Crohn’s Disease

The incidence of pediatric Crohn's Disease (CD) is increasing worldwide, particularly in countries with previously reported low rates of inflammatory bowel diseases (IBD). The dramatic rise in incidence of IBD in newly industrialized countries, shifting to western dietary habits, represents one of the foremost clues of the relevant influence of environmental factors, among which diet, in the pathogenesis of CD. Despite numerous available medications, achieving prolonged remission remains challenging for many patients, often requiring long-term immune suppression. Guidelines recommend dietary therapy for children with mild-to-moderate CD, especially exclusive enteral nutrition (EEN). EEN involves liquid medical formula as the sole energy source for 6–8 weeks with excellent remission together with many limitations that include poor adherence, social and mental-health impacts, and need for multidisciplinary support. In response to these challenges, the Crohn’s Disease Exclusion Diet (CDED) has emerged as an alternative approach. CDED is a whole-food diet specifically designed to minimize exposure to potentially inflammatory dietary components and avoid exposition to noxious substances hypothesized to have harmful effect on microbiome, intestinal permeability, and immunity. From disease monitoring point of view, the quantitative determination of the short chain fatty acids (SCFAs) in biological fluids has gained much attention in the last years because of their relevant implication as fermentation end products of gut microbiota. The fecal sample has gained a major role being the most accessible biological matrix which can directly probe the connection between intestinal bacteria and the physiology of the holobiont. SCFAs abundance depends on the amount of microbiota present in the colon, the substrate source, and the gut transit time. Alterations of SCFAs circulating concentrations reflect shifts in gut microbiota diversity and richness, classified as a dysbiosis status that has been strongly associated with IBD. The present study was aimed to evaluate a possible correlation among SCFAs, therapy and disease remission status. The content of SCFAs, in fecal samples of 17 pediatric patients affected by CD, was measured by gas chromatography coupled to mass spectrometry, using solid phase microextraction as the sample preparation technique according to a previous developed method. In total, 71 biological samples of patients who received CDED were collected longitudinally and population characteristics were recorded. All the patients included in our cohort had active disease, documented by an increase of fecal calprotectin and others biochemical parameters. Quantitative data of fecal fatty acid content (C2-C5) were studied using the multivariate principal component analysis (PCA) technique together with clinical, demographic, anthropometric and laboratory data. The impact of biological therapy on gut SCFA-producing bacteria was also analysed. PCA highlighted that the content of SCFAs was correlated to the severity of the disease.