Formation analysis of primary and secondary metabolites during angkak fermentation based on GC-TOF-MS, GC-FID, and HPLC and metabolomics analysis

Abstract

The significant primary and secondary metabolites occurred during angkak fermentation are composed of organic acids, amino acids, monacolin K, pigments, and citrinin (a mycotoxin), which their variance is able to inform how the Monascus-fermented produce characteristics. Here, gas chromatography time of flight mass spectrometry (GC-TOF-MS), gas chromatography-flame ionization detector (GC-FID), and high-performance liquid chromatography (HPLC) were utilized for analysis of the metabolites in angkak (Monascus purpureus) for 30 days. Principal component analysis (PCA) and Pearson’s correlation coefficients were to study the relationship between the metabolites and fermentation time. Moreover, the metabolomics analysis in angkak was performed by using MetaboAnalyst 5.0. The amount of primary metabolites such as the organic acids (succinic acid, glycolic acid, oxalic acid, lactic acid, lactobionic acid, and pyruvic acid), amino acids (valine, isoleucine, and alanine), fatty acids (stearic acid, oleic acid, linoleic acid, palmitic acid, and linolenic acid), and, monosaccharides (i.e. fructose and glucose) increased gradually until 15 days of the fermentation and then would decrease (the death phase of M. purpureus). For the secondary metabolites, the levels of monacolin K, Monascus pigments, and citrinin (a mycotoxin)after day 10 of the fermentation increased more than 2 times as compared with the previous fermentation time of day 5, exposing to begin the stationary phase of M. purpureus growth. Similarly, according to the heat map of primary metabolites’ production rates, there were their higher production rates of primary metabolites after 10 and 15 days of the fermentation; whereas, the higher production rates of secondary metabolites were found after 25 and 30 days of the fermentation. The PCA and Pearson’s correlation coefficients explained that the primary and secondary metabolites were discriminated completely into different groups. Based on the data of 19 metabolites throughout 30 days of the fermentation, there were estimated 23 metabolisms, but outstanding 4 metabolisms were glycolysis, pyruvate metabolism, α-linolenic acid metabolism, and linoleic acid metabolism.