Study On The Quality Standard And Stability Of TP API And Tablets

TP is a developing natural medicine belonging to the fifith type of new drugs,this article mainly studied the quality standard and stability of TP API and its tablets as the antioxidant medicine.TP contains the extracted mixed polyphenols,its quality should be controlled by monomer phenols and total polyphenols that are effective ingredients.therefore,this paper established a HPLC method for determining the content of three kinds of monomers（contentgallic acid,catechin and epicatechin）and a UV method（Folin phenol method）for determining the content of total phenol,through method validation,the HPLC method and UV method are accurate and reliable,and good repeatability,and is suitable for analysis of monomers and polyphenols.And the stability of TP API and its tablets are studied through impact factor test,accelerated test and long-term test.1 Study on the quality standard of TPAPI（1）General inspectionTP API is red brown powder,has special aroma and its flavour is acerbity.The result of solubility experiment show that its insolubles accounted for 2.45%in water solution（sample concentration is 0.3 mg/m L）,2.48%in 20%methanol aqueous solution（sample concentration is 2.0 mg/m L）;hygroscopicity creation of three groups of TP APIS all reach 10%,and chloride,sulfate,heavy metals,arsenic salts and on ignition residue,ash content examination were all less than 0.016%,0.016%,20ppm,2 ppm,3.3%,2.4%.（2）The establishment of the monomer polyphenol determination methodTo establish an HPLC method for determination of three kinds of monomer phenols（gallic acid,catechin and epicatechin）.Chromatographic conditions:chromatographic column:Agilent zobax SB-C₁₈column;Mobile phase:acetonitrile（A）-（1%methanol+0.15%TFA）water（B）;Flow rate:1.0 m L/min;detection wavelength:280 nm;injection volume:20μL;Column temperature:40℃;Sample concentration:2.0 mg/m L;Gradient:0-10 min,0%→7%A;10-30 min,7%→16%A;30-40 min,16%→90%A;40-50 min,90%A;50-55 min,90%→0%A;55-65 min,0%A.Through methodology validation,the method is accurate,sensitive,reliable,good durability,and is appropriate for determination of monomers.The determination content of gallic acid,catechin and epicatechin were 0.69%,3.15%,2.48%,total content of three monomer phenols 6.32%.（3）The establishment of the total polyphenol determination methodUsing Folin-ciocaheau method to determinate the content of polyphenols,the principle is that polyphenol in alkaline conditions,react with Folin-ciocaheau reagent to produce blue compounds,whose maximum absorption is 760 nm.Precisly moving samples 1.0 m L from TP aqueous solution（0.3 mg/m L）in 50 m L brown volumetric flask,add orderly 30 m L water,2.5 m L Folin-ciocaheau reagent,10 m L 20%sodium carbonate solution,add water to scale,shake up;placing the brown volumetric flask to dark place 2h,determining its absorbance at 760 nm.through methodology validation,the method is sensitive,reliable and reproducible.The total content of polyphenol is more than 65%.（4）The establishment of the organic residual solvent determination methodBecause TP is extracted by ethanol through macroporous resin of AB-8,so there may be residual solvents of hexane,benzene,ethanol,methylbenzene,o-xylene,m-xylene,p-xylene,styrene,1,2-diethyl benzene and so on in TP API.This paper establish a method for determining the content of above nine kinds of residual solvents by headspace-capillary gas chromatography.the chromatographic conditions as follows:the chromatographic column:DB-WAX（30m×0.45 mm,0.85μm）capillary column,carrier gas is N₂,the flow rate is 3.5 m L/min,split ratio is 2:1,injection port temperature is 220℃,detector temperature was set at 250℃.Headspace condition:furnace temperature is 80℃,transfer line temperature is95℃,loop temperature is 110℃,the sample solution balance time is 30 min.The column temperature program was initially 30℃（keep for 11 min）,increased at the rate of 30℃/min to 60℃,and increased at the rate of 5℃/min to 90℃,at the end increased at the rate of 10℃/min to 200℃（keep for 2 min）.According to the results,each component was separated relatively well,and has a good linear relationship in their respective concentration range,the recovery rate was between90%～105%on average.After the examination,there are only benzene,ethanol,styrene detected,and are all within the prescribed limits.（5）the analysis for components in TP by LC-MSThrough LC-MS analysis,there are three monomer phenols and procyanidinic mainly includes seven kinds of non-ester dimer,three kinds of ester dimer,five kinds of non-ester trimer,one kind of one-ester trimer,four kinds of non-ester type four polymers and three kind of non ester five polymers in TP extracts.Thus it prove well on the composition of TP are mainly low polymer procyanidins.2 TP tablet quality standard researchThe study mainly contain the appearance,character,identification,inspection,determination of monomer phenols and total polyphenolic content of TP tablet.Through specificity test,materials are all no interference with the determination of monomers and total polyphenol.Dissolution rate is in the condition of 900 m L water-0.5%SDS-75 r/min-37℃,take the sample solution at 45 min,dissolution amount of TP tablet（caculate with total polyphenol）must be greater than 75%.3 stability studyTo test the stability of the TP API and its tablets,placeing ten days at the condition of the influencing factors of high temperature,high humidity and strong light,and to test the stability of the active ingredients in the long-term and accelerated test.High humidity test show that TP API have increased water largely,but content of monomers and polyphenol both have no change,Results of high temperature and strong light tests show that the content of gallic acid and polyphenol had no largely change,but content of catechin and epicatechin were lower than zero day’s.TP tablets only changes in high humidity test with monomers and polyphenol both decreased because of adding water,In high temperature and strong light tests which have no change.therefore,not only API but also tablet should keep in a cool,dark and dry place.Currently long-term and accelerated testing of TP API are progressing to the nine months,tablet to the three months,As time gose on,content of monomer phenols has a slight decline,change of total polyphenol content is not obvious.The test is still in progress.