Study On The Andrographolide Extracts Using Popliteal Lymph Node Assay In Mice And P815Mast Cell Degranulation Model

Objective: Anaphylactoid reactions due to contact with andrographolide injections haverepeatedly been reported. Some possible references for quality control and safety of CXLIare needed to provide through the research the selection of sensitizations from andrographisexacts using D-PLNA.Method:(1) To established popliteal lymph node assay: female BALB/c mice wererandomly divided into several groups.mice were injected subcutaneously (50μL) with drugsinto the right pad of hind foot.animals were sacrificed on the6th day,the popliteal lymphnodes(PLNs) were isolated and weighed respectively to calculate the PLN weightindex(WI)and cell index(CI),the paraformaldehyde solution for histopathologicexamination.(2) the reliability of PLNA was proved by three kinds of clearing-heat traditionalchinese medicine injection (TCMI).(3) used both the direct popliteal lymph node assay(D-PLNA) in balb/c mice and avitro model of P815mast cell degranulation to evaluate the immuno-sensitizing potential ofextract from Andrographis paniculata Nees. weight(WI) and cellularity (CI) indices forPLNs as indexes, combined with popliteal lymph node AE dyeing pathological structurefigure,D-PLNA was performed with andrographolide injection, excipient and extracts toscreen the sensitizing component. Meanwhile the model of P815mast cell degranulationwas conducted the research on the component that had been used in D-PLNA. The histamine levels,cell degranulation percentages and ammonia glycosidase releases weremeasured to explain the potential of the sensitizing agent from andrographolide extracts.Result:(1) successfully established popliteal lymph node assay.50μgHgCl2and0.75mgDF as positive control drugs.As the result of WI and CI,HgCl2(WI=3.39±0.76,CI=6.82±1.28) and DF(WI=2.33±0.35，CI=5.51±0.93).(2)ZJFI,QKLI and CXLI were responded to positive in D-PLNA, BLGI and CHIwas negative.those results were consistent with the clinical adverse drug reactions.D-PLNAis reliable in selecting the sensitization of the drugs.(3)AEE,TAL,EAE and NBE significantly increased PLNs weight and cell number(P<0.05or P<0.01) with WI>2and CI>5(considered as positive responses), Relative to theuntreated side, NAD,AND and DDA achieved the criterion of a positive PLNresponse(WI>2and CI>5),but their aqueous extract (AE) results failed (WI<2and CI<5).In the cell model,different extract fractions and three monomers of andrographolide allmarkedly elevated the histamine level,the number of degranulated mast cell and ammoniaglycosidase releases.Conclusion: In the D-PLNA, the best time is the5th day to remove the PLN. Thepopliteal lymph node is suitable for evaluating the sensitizations of drugs.Andrographolideextract (AEE,TAL, EAE, NBE) and three paniculta lactone monomer (NAD AND, DDA)are potentially allergenic. Paniculta lactone is its anti-inflammatory, antipyretic,antibacterial active ingredients as well as sensitizations of the material. So in the qualitycontrol of andrographolide injection, on the basis of guarantee the safe and effective, as faras possible to reduce paniculta lactone content, it is beneficial to enhance security, but alsosaving the medicine. Its safe and effective dose is worth our further research.