The Research Of Quality Control Method For Biological Activity Of Lumbricus

Objective:The fibrinolytic activity of different sources was obtained by identifying the varieties of Dilong.To establish a method of bioavailability of earthworm and evaluate its fibrinolytic activity with different processing methods from the same source.Methods:1.DNA was extracted from 12 batches of samples.Sequence Amplification and Bidirectional Sequencing Stitching with CO1 Universal Primers.Four batches were sequenced by 12 S rDNA primer amplification.The sequence of related species was downloaded from Genbank and analyzed by MEGA-X software.2.The fibrinolytic activity of different processed samples from different sources was compared by fibrin plate method.Results:1.Fifteen CO1 gene sequences were obtained from 12 batches.Four 12 s gene sequences in four batches.The purchase of Daping No.2 was identified as Eisenia sinensis,Two of the three batches of samples were identified as Lumbricoides magna.One batch of samples are authentic Guangdilong.Eleven batches of mountain dragons were identified as fake.Sequence alignment of five batches of Shanghai Dilong meridians is genuine.Three batches were identified as Popular Far Blind Earthworms.Two batches were identified as William Lumbricoides2、The RSD value of thrombin in different batches was 1.71%,Linearity: The sample size ranged from 12.5 to 400 U,the regression equation Y = 0.9824 X + 97.651,R2 =0.9531.The recovery rate was 86.16%,RSD was 9.65% and RSD was 0.98% in precision test.The sample was stable at lower than 60 degrees,and the fibrinolytic activity of several batches of earthworm medicinal materials was obtained.3.The fibrinolytic bioavailability of different processing methods varied greatly from different producing areas,and the results ranged from 0 to 3122 IU/mg.The freeze-drying method had the highest value,and the freeze-drying method was superior to the traditional processing method.4.With the increase of temperature,the biological activity of Lumbricus tended to decrease,but it was stable below 60 ℃.Conclusion:1、Using CO1 and 12 S rDNA primers to amplify short DNA barcode sequence of earthworm,we can quickly and accurately identify different primordia of earthworm.2.Fibrin plate method is fast,accurate,high throughput,easy to operate and reproducible.It can be used as one of the evaluation methods of fibrinolytic bioactivity of earthworm.3、The fibrinolytic bioavailability of earthworm varies greatly from place to place and from different processing methods.