Preparation Of Metarhizium Anisopliae And Berberine And Their Wettable Powder And Water Dispersible Granuie

Belonging to Metarhizium genus,Metarhizium anisopliae is a broad-spectrum insect parasitic fungus.This fungus can infect more than 200 species and genera of insects worldwide.M.anisopliae not only shows strong pathogenicity and substantial insecticidal effect,but also is non-toxic to human,livestock and crops and has broad applications.The process of infecting insects includes adhesion,spore germination,penetration,blood development and death.Its main independent unit is aerial conidia.Berberine is a quaternary ammonium isoquinoline alkaloid of berberine base isolated from Coptis chinensis.Berberine has many clinical effects,such as anti-inflammation,antioxidation,protection of neuro cardiovascular system,treatment of chronic heart disease,attenuation of lipid,improvement of insulin resistance,etc.Moreover,its broad-spectrum bactericidal effect makes it a biological pesticide applied in agriculture and animal husbandry.Through reviewing literature and experiment,it was found that M.anisopliae was not sensitive to berberine.Based on the quality index of the original medicine of metaspore powder of M.anisopliae CQMa421,the EC50 of berberine against M.anisopliae was determined.By summarizing the field experiment results of M.anisopliae and berberine,the mixture ratio of berberine and M.anisopliae was selected.The compatibility of carriers and additives was selected,and the formulation optimization and quality test of M.anisopliae and berberine wettable powder were carried out.Furthermore,based on development of wettable powder,the water dispersing agent was developed by extrusion granulation and the quality index was tested.The main results are summarized as follows:1.Production and quality index detection of M.anisopliae spore powderRice was sterilized by high pressure for 9-11 days,dried at 30-37℃for 3 days.The material was roughly grinded and then passed through an 80 mesh cell sieve and stored at low temperature and dark.The spore content of M.anisopliae conidia was 4.26 ×1010 CFU/g.After dried to constant weight,the average moisture was found to be 2.39%,and the living spore rate was 95.02%.2.Preparation and quality control of compound wettable powder of M.anisopliae and berberineSince berberine has antibacterial activity,its biocompatibility with M.anisopliae should be determined prior to their compounding.Generally,the larger the EC50 is,the safer the examined sample is.It was shown that the EC50 of berberine to M.anisopliae was 47.2 mg/mL.Compared with the EC50 value(8.314 μg/mL)reported for other pathogenic bacteria by our laboratory,the conidial growth was not affected by the same concentration of berberine.Taking the field experiment of M.anisopliae into account,the content of berberine was determined as 2.66%-3.54%.Taking wettability test into account,the proportion of spore powder was determined as 25%.First,we screened the carrier,wetting agent,dispersant,protective agent and other additives by biocompatibility test.Next,the additives were re-screened.Taken together,the formulation of wettable powder was determined as follows:M.anisopliae spore powder of 25%,berberine content of 2.66%-3.54%,wetting agent NP-10 of 5%,dispersant MF of 2.05%and calcium lignosulfonate of 5.12%,protective agent L-ascorbic acid of 0.5%,carrier diatomite supplement to 100%.The quality test showed that the living spore rate of the preparation was 83.4%,the water content was less than or 1.63%,the wetting time was 58s,the spore suspension rate was 81.34%,the fineness(passing through 325 mesh cell sieve)≥ 99%,and the pH value was 5.5-6.2.The performance index meets the standard.3.Preparation and quality control of M.anisopliae-berberine compound Water dispersible granulOn the basis of the second chapter,we continued to select disintegrant and binder types.To this end,response surface methodology was used to optimize the constitution of dispersant,disintegrant and binder.As a result,the formula was determined as follows:M.anisopliae spore powder of 25%,berberine content of 2.66%-3.54%,wetting agent BX of 5%,dispersant MF 2.1%,calcium lignosulfonate 5.3%,protective agent L-ascorbic acid of 0.3%,disintegrating agent sodium alginate of 18%,binder sodium carboxymethyl cellulose of 1.7%,and carrier supplement of 100%.The quality test results showed that disintegration time was 176s,live spore rate was 80.4%,and spore suspension rate was 86.34%.