Development and Validation of a LC-MS/MS Technique for the Analysis of Short Chain Fatty Acids in Tissues and Biological Fluids without Derivatisation Using Isotope Labelled Internal Standards

Abstract

The gut microbiota is critical to the maintenance of physiological homeostasis and as such is implicated in a range of diseases such as colon cancer, ulcerative colitis, diabetes, cardiovascular diseases, and neurodegenerative diseases. Short chain fatty acids (SCFAs) are key metabolites produced by the gut microbiota from the fermentation of dietary fibre. Here we present a novel, sensitive, and direct LC-MS/MS technique using isotopically labelled internal standards without derivatisation for the analysis of SCFAs in different biological matrices. The technique has significant advantages over the current widely used techniques based on sample derivatization and GC-MS analysis, including fast and simple sample preparation and short LC runtime (10 min). The technique is specific and sensitive for the quantification of acetate, butyrate, isobutyrate, isovalerate, lactate, propionate and valerate. The limits of detection were all 0.001 mM except for acetate which was 0.003 mM. The calibration curves for all the analytes were linear with correlation coefficients r2 > 0.998. The intra- and inter-day precisions in three levels of known concentrations were <12% and <20%, respectively. The quantification accuracy ranged from 92% to 120%. The technique reported here offers a valuable analytical tool for use in studies of SCFA production in the gut and their distribution to host tissues.

A Chiral-LC-MS Method for the Simultaneous Quantification of Short-Chain Fatty Acids and D/L-Lactate in the Ruminal Fluid of Dairy Cows

“…Compared with non-derivatised SCFA [ 21 ], our method shows greater sensitivity and selectivity and affords better peak shape. Due to the high abundance of SCFA and D/L-lactate in ruminal fluid, an LOD of 0.01 µg/mL is more than adequate for such samples, so no attempt was made to further improve the MS response (e.g., using a narrower column and reducing the elution flowrate).…”

Anaerobic phloroglucinol degradation by Clostridium scatologenes

“…A gradient of 0–4 min 0%−60% B, 4–8 min 60%–100% B, and 8–12 min 100% B was used. The mass spectrometry detection was performed under positive electrospray ionization mode [ESI (+)], and the analytes were monitored in multiple-reaction monitoring mode ( 57 ) .…”

Determination of short-chain fatty acids as putative biomarkers of cancer diseases by modern analytical strategies and tools: a review

“…The gold-standard method such as colonoscopy, biopsy, or use of contrast labels are screening techniques with high invasiveness and costs. According to novel studies, different types of carcinomas could be detected by SCFAs in the fecal, serum, plasma, tissues or semi-skimmed milk samples as oncological markers by analytical methods with high sensitivity and precision ( 47 , 48 ). An excellent example is a colorectal carcinoma where the implementation of custom-made methods with minimal invasiveness, protection, and reproducibility are highly desirable.…”