ABSTRACT
A high-content GABA was found in Sojae Semen Praeparatum(SSP), which is a famous traditional Chinese medicine and officially listed in Chinese Pharmacopoeia. To screen out and identify GABA-producing microbes from samples at different time points during the fermenting process of SSP, traditional microbiological methods combined with molecular biological methods were used to study the predominant GABA-producing microorganisms existing in the fermenting process of SSP. This study would lay a foundation for further studying the processing mechanism of SSP. The fermenting process of SSP was based on Chinese Pharmacopoeia(2010 edition), and samples were taken at different time points during the fermenting process of SSP. The bacteria and fungi from samples at different time points in the fermenting process of SSP were cultured, isolated and purified by selective medium, and dominant strains were selected. The dominant bacteria were cultured in the designated liquid medium to prepare the fermentation broths, and GABA in the fermentation broth was qualitatively screened out by thin-layer chromatography. The microbial fermentation broth with GABA spots in the primary screening was quantitatively detected by online pre-column derivatization and high performance liquid chromatography established in our laboratory. GABA-producing microorganisms were screened out from predominant strains, and their GABA contents in fermentation broth were determined. The DNA sequences of GABA-producing bacteria and fungi were amplified using 16S rDNA and 18S rDNA sequences by PCR respectively. The amplified products were sequenced, and the sequencing results were identified through NCBI homology comparison. Molecular biological identification was made by phylogenetic tree constructed by MEGA 7.0 software. Through the homology comparison of NCBI and the construction of phylogenetic tree by MEGA 7.0 software, nine GABA-producing microorganisms were screened out and identified in this study. They were Bacillus subtilis, Enterococcus faecium, E. avium, Aspergillus tamarii, A. flavus, A. niger, Cladosporium tenuissimum, Penicillium citrinum and Phanerochaete sordida respectively. For the first time, nine GABA-producing microorganisms were screened out and identified in the samples at different time points during the fermenting process of SSP in this study. The results indicated that multiple predominant GABA-producing microorganisms exist in the fermenting process of SSP and may play an important role in the formation of GABA.
Subject(s)
Bacteria , Classification , Metabolism , Chromatography, High Pressure Liquid , Fermentation , Fungi , Classification , Metabolism , Phylogeny , Seeds , Microbiology , Glycine max , Microbiology , gamma-Aminobutyric AcidABSTRACT
OBJECTIVE: To investigate the anticonvulsion effect of 4-amino-2-methyl cantharidinmide (AMC) and its influences on epileptiform electroencephalogram (EcoG), contents of γ-aminobutyric acid (GABA) and GABAB receptor. METHODS: Rat model of penicillin-induced-convulsion (PIC) was established by intracortical (ic) penicillin (PNC) injection in rat motor cortex. Valproate (VPA) was used as the positive control drug. Convulsion seizure latency and racine behavior study graduations were used as indexes to evaluate the efficacy. RM6240C multi-channel biological signal collection-processing system synchronously recorded EcoG of convulsive rats after intragastric (ig) administration of AMC (25.0, 100.0 mg · kg-1). The effects on convulsion and epileptiform discharge were analyzed. The contents of GABA and the expression of GABAB receptor in the cortex and hippocampus regions of rats were determined by immunohistochemistry technique. RESULTS: AMC at the two doses and VPA could reduce epileptiform activities and discharge and prolong the latencies of epilepsy seizure, compared with the PIC group (P 0.05); compared with model control, the expression quantity of hippocampal GABAB receptor protein was significantly increased (P 0.05). GABA expression quantity in groups of larger dosage AMC and VPA were respectively higher than that in the normal group and the model group, and the GABAB receptor protein expression quantity also significantly increased (P < 0.05). CONCLUSION AMC can antagonize the convulsion seizure and inhibit the epileptiform discharge induced by penicillin in rats. The anti-convulsion mechanism of AMC is possibly related with increasing GABA content and GABAB receptor expression in cortex and the hippocampus. Copyright 2012 by the Chinese Pharmaceutical Association.
ABSTRACT
According to 5 different subtypes of γ-amino butyric acid transporter(GAT)in brain regions and subcellular distribution.GAT1 and GAT3 are closely related with occurrence and development of epilepsy.The abnormal expression of GAT or their function damaged contribute to hyperexcitable neurons In seizures.GABAergic inhibit circuit reduces and the down-regulation of GAT expression,primary up-regulation of GAT expression is a reactive changes or cause of epilepsy.
