| ObjectiveThe formulation of pharmaceutical stoppers is complex and includes rubber materials and a variety of rubber ingredient(e.g.vulcanizing agents,vulcanization accelerators,antioxidants,plasticisers,etc.).Throughout the production process,’vulcanization’is an indispensable step,which crosslinks the linear molecular structure of the rubber into a three-dimensional mesh structure through high temperatures,enhancing the product’s performance.This process requires the involvement of a vulcanizing system,in which the use of vulcanizing agents and vulcanization accelerators plays a key role.However,studies have found that there are safety risks associated with the addition of these two types of rubber ingredient.Firstly,they are toxic themselves or decompose to produce carcinogenic substances(N-nitrosamines,aniline)residues,and secondly,they may migrate or even react chemically with the drug over a long period of time,reducing the efficacy and threatening human health in serious cases.Therefore,a multi-level,cross-applicable quality control and safety evaluation system for the vulcanization of pharmaceutical stoppers with vulcanizing agents and vulcanization accelerators was developed using triple chromatographic analysis techniques.MethodsIn this paper,the quality control and safety evaluation system of the vulcanizing agents and vulcanization accelerators were studied based on three analytical techniques,using products with different vulcanization systems and different rubber varieties produced by four pharmaceutical stopper manufacturers in different regions of China.The main methods are as follows:1.Study of high performance liquid chromatographic methods for the analysis of 2vulcanizing agents and 12 vulcanization accelerators in pharmaceutical stoppers with five types of vulcanization systems.2.Study of Gas chromatographic-sulfur chemiluminescence detector for the analysis of vulcanizing agents and vulcanization accelerators in pharmaceutical stoppers.2.1 Determination of 9 vulcanizing agents and vulcanization accelerators in pharmaceutical stoppers by headspace gas chromatography-sulfur chemiluminescence detector(HS-GC-SCD).2.2 Drug compatibility-migration test.3.Study of a heart-cutting/two dimensional liquid chromatography-tandem mass spectrometry(2DLC-MS)method for the analysis of vulcanizing agents and vulcanization accelerators in pharmaceutical stoppers.3.1 Simultaneous determination of 3 vulcanizing agents and 13 accelerators in pharmaceutical stoppers of different vulcanization systems based on heart-cutting/two dimensional liquid chromatography-tandem triple quadrupole mass spectrometry.3.2 Drug compatibility-migration test.Results1.For the analysis by HPLC,a Waters Sun Fire C18 column was selected and methanol-water solution was used as the mobile phase;after determining the chromatographic conditions,the qualitative and quantitative analyses of the 2vulcanizing agents and 12 vulcanization accelerator standards were carried out.The results showed that the linearity of these 14 compounds was good in the concentration ranges of 0.10～20,0.50～20 and 1.0～20μg/m L,respectively,with the correlation coefficients greater than 0.9950;the relative standard deviations(RSDs)of precision,reproducibility and stability were less than 2%.The average recoveries of the samples from different sulphuration systems ranged from 90.0%to 108.7%,and the RSDs ranged from 0.10%to 2.8%.From the results,the presence of sulphur was detected in most of the samples at 0.0030～0.2114 mg/g;N-Cyclohexyl-2-benzothiazole hypersulphonamide was detected in three batches at 0.0708～0.4312 mg/g.2.Establishment and methodological study of the HS-GC-SCD method.An Agilent DB-1 capillary column was selected with a size of(30m×0.32mm,0.25μm).The headspace conditions and chromatographic conditions were optimized to qualify the retention time of the standards and quantify them by external standard method.9sulphides were linear in the concentration ranges of 0.5～20μg/m L and 2.0～50μg/m L with the correlation coefficients greater than 0.9970;the RSDs of precision and reproducibility were 0.59%～2.70%and 0.17%～1.99%,respectively.The average recoveries were 82.8%～99.0%.Only free sulphur was detected in the samples at0.010～11.647μg/g,and the remaining components were not detected.In addition,the results of the compatibility study tests showed that none of the nine sulphurising agents and accelerators migrated.3.A heart-cut/two dimensional liquid chromatography-tandem mass spectrometry analysis system was constructed.The content was determined in positive ion mode with electrospray ionization(ESI~+)and dynamic multiple reaction monitoring(DMR).3 vulcanizing agents and 13 vulcanization accelerators showed good linearity in the corresponding mass concentration ranges with correlation coefficients greater than0.9900 and the LODs were 0.12～5.0μg/kg;the average recoveries were90.0%～107.0%and the RSDs were 0.50%～2.9%.The results showed that di-tert-butyl peroxide was detected in 4 batches with the detection amounts of15.99～23.68 mg/kg;N,N’-dicyclohexyl-2-benzothiazole hypersulfonamide was detected in 4 batches with the detection amounts of 43.07～77.37 mg/kg;N-tert-butyl-2-benzothiazole hypersulfonamide was detected in 12 batches with the detection amounts of 0.11～3.19 mg/kg.Meanwhile,the results of the compatibility study indicated that all the vulcanizing agents and accelerators were not detected in the migration amount.ConclusionIn this thesis,a system for the quality control and safety evaluation of vulcanizing agents and accelerators in pharmaceutical stoppers of different vulcanisation systems is systematically developed for different scenarios.The three analytical methods established:HPLC method is widely used and popular,GC-SCD method is more specific for sulphides,and 2DLC-MS method can be used for precise qualitative and quantitative analysis.The analysis of a total of 20 vulcanizing agents and accelerators in three commonly used systems has been achieved.This is the first time that SCD and two-dimensional liquid-phase techniques have been applied to the study of the residues of vulcanizing agents and accelerators in pharmaceutical stoppers and their migration into drugs,laying a solid foundation for the study and analysis of vulcanization systems in pharmaceutical stoppers.In addition,the migration test study of the examined stoppers with simulated packaging of sodium chloride injection suggested that the migration of the vulcanizing agent and accelerator studied in this paper did not occur in the simulated drug,which can be considered as a low safety risk and the drug compatibility of the medicinal stoppers is good.It also indicates that the quality control and safety evaluation system of the vulcanizing agent and accelerator in the pharmaceutical stopper can be effectively applied to the study of the compatibility of the pharmaceutical stopper with pharmaceuticals. |
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