Taxonomy And Identification Of The Genus Scolopendra And Quality Evaluation Of Centipede Medicinal Materials In China

The genus Scolopendra are widely distributed around the world,especially in tropical and subtropical areas.As an ancient biological species,it is an important branch of arthropod evolution,and its study of taxonomy has great significance to species evolution.Due to limited mobility,different Scolopendra species live in relatively independent geographical distribution regions,which make them become important historical geography research material.Scholars have explored the taxonomy of this genus,but the status of some species are controversial,especially at the subspecies of S.subspinipes subspinipes and S.morsitans.The relationship between species is still uncertain.In China,the species of Scolopendra are mainly distributed in the south of qinling-huaihe isotherm.However,the depth research on resource of genus Scolopendra in China is rarely conducted,and the number and distribution of these species in China are not clear.Morphological identification is a common method for taxonomy and identification of the genus Scolopendra,but the similarity in characteristics often make some individuals overlapped in morphology,and when some parts are absent or samples become powder,it is difficult to identify these species.In recent years,other methods,such as geometric measurement or molecular biology,have been raised to apply for the taxonomy and identification of the genus Scolopendra,and acquired certain breakthrough,but the bottleneck is still not solved by single method.More methods were needed to systematically study the taxonomy of the genus Scolopendra.As traditional Chinese medicinal material,medicinal centipedes,which are mainly from the genus Scolopendra,has been used for more than 2000 years.Recently,with the in-depth of research on its efficacy,anti-tumor,analgesic,anti-inflammatory and other effects have been continuously found,and the clinical application has been gradually expanded,which make medicinal centipedes become one of the hotspots of research.S.mutilans is the only species which recorded by Chinese pharmacopoeia,but some other species are also used in some regions in China.The species of Scolopendra are toxic animals,and the toxicity and efficacy are different among species.However,there are confusion and misuse among species due to the similarity of characteristics,which causing potential harm to human health.Accurate identification of these species will be helpful for solve the confusion and misuse problems.The centipede medicinal material is from animal,with unclear effective ingredient and simple quality control methods,which is difficult to evaluate the quality.The study of multi-indicator quality control method of centipede medicinal material will be helpful to improve the quality standard system and ensure the safety and quality.In this study,a systematic study of the genus Scolopendra in China was carried out,which provided a new idea for the resource,taxonomy and quality evaluation of Chinese medicinal centipedes.The main contents are as follows:1.The resource research of the genus Scolopendra in China was carried out through visiting,sampling in field and investigating from medicinal materials market or company.Combined with previous reports,the resource and distribution were studied,14 species of the genus Scolopendra were determined to live in China,and "three river systemic zones" and "three geographic areas" was put forward for the distribution of medicinal centipedes.2.The combined method of morphological-molecular was applied to classify and identify species of the genus Scolopendra.8 species were identified using morphological characteristics,and the identification key of 14 species of the genus Scolopendra in China was established.Four typical characteristics of samples were measured with morphometric method,and the statistical results were used to analyze morphological difference among species.Based on the COI gene sequences of DNA barcode technology,the phylogenetic trees of 12 nominal species were built,10 species were separated as independent branches,while S.subspinipes subspinipes and S.hainanum clustered into the same branch.Through this comprehensive method,the species,such as S.multidens and S.mutilans,etc,were determined as separated species,while S.subspinipes subspinipes and S.hainanum were treated as two "morphological-species" for their consistency of molecular data,and the samples collected from LuFeng,Yunnan,were identified as a new species and named as S.lufengensis sp.nov..3.Methods of quality control on Chinese centipede medicinal materials were studied,and the quality standard system was improved.In the qualitative methods,the characteristics differences among species were compared through morphological identification,and the microcharacter of head plates and antennae of 6 species were observed using electron microscopy the first time.After the DNA molecular method was established using DNA barcode technology,each species was clustered into a single branch in the NJ tree.Based on PCA-SVC algorithm,the NIRS rapid identification model was also explored,the results of accuracy rate were 100% in training set and 81.82% in test set,which met to the requirement of rapid identification.In the quantitative analysis,the total nitrogen contents of different species were determined by semi-micro determination method,and based on the nitrogen contents,the PLS-SVM model was built and determined as the best NIRS quantitative method for Chinese medicinal centipedes.The results showed that the qualitative and quantitative methods as above can be used to control the quality of medicinal centipedes effectively,and the quality can also be evaluated.4.The safety of Chinese medicinal centipedes was evaluated by the preliminary study on endogenous toxicity and exogenous harmful substance.The toxic ingredients of venom and medicinal materials were analyzed,and the toxicity of centipede medicinal materials to human gastric cancer cell MGC803 was investigated by MTT method.The results showed that the toxicity has potential harm to human body,the rationality of preparation could reduce the hazard risk.The residue of heavy metal elements and aflatoxin in centipede medicinal materials were studied.When the contents of five kinds of heavy metal element were determined using ICP-MS,the contents of Cu element in 75% sample exceeded standard,which generally have higher value,while Cd element in 45% sample exceeded.After the content of aflatoxin was assayed by LC-MS method,the aflatoxin was not detected out in samples,except one batch of 0.9 ?g/kg,and the aflatoxin in all samples was not exceeded standard.The results suggested that medicinal centipedes,in their growth environment,were easily loaded elements of Cu and Cd,which affecting human safety.When the storage conditions controlled strictly,the contamination of aflatoxin could be reduced.The control of heavy metal elements and aflatoxin could improve safety of medicinal centipede.In our research,the resource of the genus Scolopendra in China was explored,14 species were determined to live in China using combined method of morphology and molecular data,a new species of the genus Scolopendra was discovered and named as S.lufengensis sp.nov..The quality control and evaluation method of medicinal centipedes could prevent confusion and misuse of species,and it could be used to improve the quality of medicinal material.The study on toxicity of medicinal material could effectively reduce harmful risks to human body and improve the safety when medicinal centipedes were used.This study will play an important role in ensuring quality and clinical drug safety of centipede medicinal material,protecting resource and sustainable utilization of the genus Scolopendra.The combination method of morphology and molecular data for taxonomy and identification of the genus Scolopendra and the quality evaluation method for centipede medicinal materials in China will provide foundation for the quality control of other animal medicines.