| Objective:The impurities in albendazole feedstock were analyzed and identified by high performance liquid chromatography-ion trap mass spectrometry(LC--Orbitrap),and the overall toxicological parameters were evaluated by ADMET Predictor 8.5 software,so as to obtain the analysis and prediction of adverse reactions.Reversed phase high performance liquid chromatography(RP-HPLC)was used to determine the impurities in albendazole feedstock and the impurities that destroyed the experiment so as to fully understand the distribution and source of impurities.The residual solvent in albendazole raw material was determined by headspace GC to understand whether the residual solvent in the process met the standard.After dry treatment and wet treatment,albendazole API was determined by ICP-oes inductively coupled plasma chromatography to understand whether the inorganic metal impurities in the process met the standard,so as to further improve the quality standard of the drug.Methods:Structure and toxicity analysis of impurities in albendazole raw materials:High performance liquid chromatography-ion trap mass spectrometry(LC--Orbitrap)was used,and positive ion file of mode ESI ion source was selected.Ulultimate XB-C18 Welch(4.6mm*250mm 2.7μm)was used.The mobile phase was 0.01mol/L ammonium acetate(pH=2.8):methanol(60:40)at a flow rate of 1.0 mL/min.The column temperature was 40℃.The first and second stage full-scan mass spectrometry were used to obtain the precise information of the elemental information,molecular mass information and fragment ion information respectively,so as to achieve the purpose of identifying the structure of unknown impurities.The ADMET Predictor 8.5 toxicological parameters were used to comprehensively predict the structural properties of the impurities.(2)Measurement of the content of related impurities in albendazole raw materials;The measuring instruments and conditions were as follows:RP-HPLC;Waters Cortecs 2.7μm 4.6*150 column;0.01mol/L ammonium acetate,using formic acid to adjust its pH to 2.8,that is,mobile phase A;Methanol was mobile phase B liquid,and the gradient elution was carried out at the flow rate of 0.7 mL/min.245nm wavelength,40℃ column temperature conditions.(3)Determination of residual solvents in the raw material albendazole:Headspace GC method was adopted.Headspace temper-ature was 80℃,headspace time was 10 min,chromatography column with 94%dimethylpolysiloxane--6%nitrile-propyl phenyl was used as the stationary phase,and temperature measurement,column temperature,injection temperature and starting temperature were set at 250℃,35℃,200℃ and 40℃,respectively.DB-624-30 m×1.8×0.32 mm(including m;After initial temperature treatment for 5 minutes,it is heated to 150℃ at a rate of 15℃ per minute and kept for 10 minutes,and then heated to 200℃ at a rate of 30℃ per minute.The split ratio is set at 20:1.The head-empty bottle is balanced at 80℃ for 10 minutes.(4)The determination of ion chromatography using ICP-OES inductive coupling,etc.:parameters were set as:1.2L/min,auxiliary gas;14L/min,plasma gas;1.2KW high frequency power;Carrier gas:high purity argon 0.7L/min;Exposure time:10 seconds;Solvent cleaning:30 seconds;Wash the sample for 90 seconds.Results:(1)A total of 13 impurities were detected,among which the molecular structure of one unknown impurity was deduced.Other impurities in the liver toxicity,acute toxicity and ca-diac toxicity are probably does not exist,and impurity E possible carcinogenic toxicity,the possibility of chromosome variation,reproductive toxicity,impurity B,C,D,G,I have exist the possibility of chromosome variation,reproductive toxicity,impurities A,F,H,K,L have reproductive toxicity may,Impurity J may have chromosomal mutation toxicity,but these impurities may not cause mutation risk.(2)The linear range of impurities A,B,C,D,E,F,G,H,I,G,K,L was 0.5-10μg/mL(r>0.999)by HPLC method.All the impurities and the main peak could be separated effectively.The average recoveries of impurities were 100.70%,99.74%,100.04%,100.39%,99.35%,100.11%,99.86%,99.66%,99.94%,100.01%,99.97%,99.93%,respectively.(3)By headspace GC method,the linear relationship of impurity methanol was in the range of 14.28～128.59μg/mL,the detection limit was 0.38592μg/mL,the correlation coefficient R value was 0.9976,and the average recovery was 98%.The linear relationship of n-propanol was good in the range of 20.4～183.6μg/mL,with a detection limit of 6.20965μg/mL,a correlation coefficient R value of 0.9967 and an average recovery of 107%.The linear relationship of impurity ethanol was in the range of 20.08～180.72μg/mL,with the detection limit of 5.12866μg/mL,the correlation coefficient R value of 0.9977 and the average recovery of 102%.The linearity of toluene was in the range of 1.94～17.46μg/mL,with a detection limit of 0.31175μg/mL,a correlation coefficient R value of 0.999 and an average recovery of 109%.The linear range of impurity acetone was 20.28-182.52μg/mL,with a detection limit of 1.10955μg/mL,a correlation coefficient R value of 0.9985 and an average recovery of 86%.(4)Using ICP-OES inductively coupled plasma chromatography,the linear relationship between the two heavy metals was in the range of 0.002～10 μg/mL,and the value of phase relationship was 0.9999.By comparing the contents of Fe and Si in different samples treated by different methods,it was found that the difference between wet and dry methods was small,with the average Fe of 18μg/g and Si of 65.52μg/g.Conclusion:In this paper,a new method was established for the determination of impurities,residual solvents and inorganic metals in albendazole API,and the chemical structures of 13 impurities were preliminarily identified.Most of the impurities are found to have toxicity risk by prediction.Finally,the quality standard of albendazole API in line with the requirements of modern analytical methodology is formulated,which provides a favorable scientific theoretical basis for improving the quality of albendazole API. |
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