Résumé
ObjectiveIn order to ensure the safety and effectiveness of clinical drug use ， the identification method of mixed and adulterated specific polymerase chain reaction （PCR） identification of Pheretima aspergillum and its processed products was established. MethodBased on the cytochrome C oxidase subunit I sequence of P. aspergillum， primers were designed to cover the whole sequences， and the stable DNA ranges suitable for the identification of Pheretima （P. aspergillum） formula granule were screened out. Specific primers were designed according to the specific single nucleotide polymorphisms （SNP） of P. aspergillum in the stable DNA range. The P. aspergillum and its mixture were collected respectively， the PCR reaction system was established and optimized， and PCR reaction system and procedure were optimized， and the tolerance and applicability were investigated. ResultWhen the annealing temperature was 62 ℃ and the cycle number was 36， both P. aspergillum formula granule and its formula particles could amplify a single specific identification band of about 170 bp， and the other 20 adulterants and negative controls had no band. ConclusionThe allele-specific PCR identification method established in this study can quickly and accurately identify the P. aspergillum formula granule. The orgin of Chinese herbal medicine and decoction pieces and P. aspergillum were accurately identified. It can also provide a reference for other studies on the quality standard research of other Chinese herbal formula granule.
Résumé
Objective Our purpose is to explore the occurrence and distribution of glutamate and metabotropic glutamate receptor in earthworm, P. aspergillum and to analyze their function significance. Methods By employing immunocytochemistry tchnique, the positive cells of glutamate and metabotropic glutamate receptor, mGluR1, mGluR2/3, mGluR4 in earthworm, P. aspergillum were investigated under a light microscope. Results It was found that glutamate positive cells existed in cerebroganglion, subpharyngeal ganglion, ventral ganglion, neurons between muscle cells,intestinal epithelium and epidermis, that thin and dense glutamate positive fibers distributed in pharyngeal nerve cycle, subpharyngeal ganglion, ventral neurochain, that mGluR1 or mGluR4 positive neurons were detected in cerebroganglion and mGluR2/3 positive neurons weren't observed in earthworm. Conclusion During glutamate positive neurons, some possess remarkable processes and matured appearance from which glutamate positive fibers in pharyngeal nerve cycle, subpharyngeal ganglion, ventral neurochain derive, another have few branches, unmatured appearance in which glutamate doesn't serve as neurotransmitter acting to mGluR1 or mGluR4 of neurons in earthworm cerebroganglion.