A method for rapid analysis of polysorbate 80 components / 中国药科大学学报

@#Due to the complex components of polysorbate 80, analysis is time-consuming and labor-intensive, so there is an urgent need to find a method for rapid analysis of polysorbate 80 components.In this study, 10 batches of samples collected from 3 domestic and foreign enterprises were analyzed by UHPLC-HRMS, with the being further results were analyzed by the ExcipientProfiler software and supplemented by the extended database.The results showed that the ExcipientProfiler software could quickly identify the [M+Na]+ peak in the mass spectrogram, and obtain the information of component distribution, the numbers of components and the degree of polymerization of the sample.Meanwhile, the numbers of components obtained by the ExcipientProfiler software could be used to distinguish the injection grade samples from the ordinary grade samples by systematic clustering analysis.In addition, it was found through further supplement that the sample contained other fatty acid ester components by manually searching the relevant extended database.The polyoxyethylene sorbitan tetraoleate components were found in the sample according to the analysis of mass spectrum data.Therefore, although this method is fast and simple, it is necessary to add polyoxyethylene sorbitan tetraoleate components and other fatty acid ester components to further supplement the information in the ExcipientProfiler software, so that it can be better used for the analysis of polysorbate 80.

Control strategy for the limit of aldehydes in polysorbates based on genotoxicity and influence on the aggregation of monoclonal antibody / 中国药科大学学报

@#An HPLC pre-column derivatization detection method was established to detect and analyze the formaldehyde and acetaldehyde in polysorbate 80 and polysorbate 20 from different manufacturers.The effects of aldehyde and acetaldehyde on the aggregation of adalimumab under different conditions were monitored.Based on the control of genotoxic impurities and the influence on the stability of monoclonal antibody preparations, the control limits of the two chemicals were preliminarily obtained.2, 4-dinitrophenylhydrazine (2, 4-DNPH) was applied as the derivatization reagent in HPLC pre-column derivatization; acetonitrile and water were used as mobile phase to perform a gradient elution on a C8 (4.6 mm × 150 mm, 5 μm) chromatographic column.The detection wavelength was 360 nm, and the external standard method was used for quantification.Verification results showed that the method was suitable for the quantitative analysis of trace formaldehyde and acetaldehyde in polysorbate 80 and polysorbate 20 . The detection and analysis of formaldehyde or acetaldehyde in different batches of polysorbate 80 and polysorbate 20 from different manufacturers showed that the content of formaldehyde and acetaldehyde were quite different. The content of formaldehyde and acetaldehyde in polysorbate 80 were significantly higher than those of polysorbate 20. After monitoring the changes of adalimumab aggregates treated by formaldehyde and acetaldehyde by size exclusion chromatography (SEC), it was found that the effect of formaldehyde on adalimumab aggregation was significantly higher than that of acetaldehyde.According to the requirements of ICH M7 (International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use, M7: Assessment and Control of DNA Reactive (Mutagenic) Impurities in Pharmaceuticals to Limit Potential Carcinogenic Risk), the impurity limits of formaldehyde and acetaldehyde in polysorbate 80 and polysorbate 20 for monoclonal antibody preparations were calculated from the perspective of risk assessment.Combined with the influence on the aggregation stability of monoclonal antibodies, the preliminary limis for acetaldehyde and acetaldehyde were recommended to be ≤ 7 μg/g and ≤ 765 μg/g, respectively.

Interlaboratory method validation of HPLC-FMA for determination of polysorbate 80 in monoclonal antibodies / 药学学报

The high performance liquid chromatography-fluorescence micelle assay (HPLC-FMA) method for the content determination of polysorbate 80 in monoclonal antibody drugs was validated to study its applicability and transferability between various laboratories, and the feasibility to be included in the Chinese Pharmacopoeia. Both J.T. Baker and Nanjing Well-sourced polysorbate 80 was used in the collaborative validation of polysorbate 80 content analysis in seven different laboratories. The results show that when the protein concentration was no more than 20 mg·mL-1 and the concentration of polysorbate 80 ranged from 0.05 to 0.5 mg·mL-1, the method had good specificity. The recovery rates of the spiked samples ranged from 92.20% to 117.70% for J.T.Baker and from 93.90% to 117.20% for Nanjing Well. The intra-laboratory precision (%RSD) was less than 4.30% for J.T. Baker, and less than 2.60% for Nanjing Well, while the overall precision was less than 5.45% for J.T. Baker, and less than 6.70% for Nanjing Well. The linear correlation coefficient was more than 0.98 for J.T. Baker and more than 0.99 for Nanjing Well. The results of the collaborative validation prove that the HPLC-FMA method has good accuracy, precision, linearity, and specificity, and could be used for release control analysis of polysorbate 80 content in monoclonal antibodies across different laboratories.

Nanosuspension enhances dissolution rate and oral bioavailability of Terminalia arjuna bark extract in vivo and in vitro

Objective: To enhance the dissolution rate and oral bioavailability of Terminalia arjuna bark extract by formulating its nanosuspension. Methods: Nanoprecipitation approach was used for the formulation of nanosuspension using polysorbate-80 as a stabilizer. The formulated nanosuspension was assessed for particle size, polydispersity index, zeta potential value and for in vitro dissolution study. Oral bioavailability studies were carried out in Wistar male albino rats by administering a single dose (50 mg/kg. b. wt) of the formulated nanosuspension and coarse suspension. The storage stability of the formulated nanosuspension was determined after three months of storage at room temperature and under the refrigerated condition. Mutagenicity assay was carried out to evaluate the toxicity of the formulated nanosuspension using two mutant strains (Salmonella typhimurium TA100 and Salmonella typhimurium TA98). Results: The mean particle size of the formulated nanosuspension was 90.53 nm with polydispersity index and zeta potential values of 0.175 and-15.7 mV, respectively. Terminalia arjuna nanosuspension showed improved dissolution rate and 1.33 fold higher oral bioavailability than its coarse suspension. The formulated nanosuspension also showed better stability under the refrigerated condition and was non-mutagenic against both strains. Conclusions: Our study demonstrates that nanosuspension technology can effectively enhance the dissolution rate and oral bioavailability of Terminalia arjuna bark extract. Zafar Fatiqa 1 Department of Chemistry, University of Okara, Okara Jahan Nazish 2 Department of Chemistry, University of Agriculture, Faisalabad Khalil-Ur-Rahman 3 Department of Biochemistry, University of Agriculture, Faisalabad Asi Muhammad 4 Food Toxicology Lab, Plant Protection Division, Nuclear Institute for Agriculture and Biology, Faisalabad Zafar Waseeq-Ul-Islam 5 Department of Computer Science, COMSATS University of Information and Technology, Islamabad Pawar SS, Dahifale BR, Nagargoje SP, Shendge RS. Nanosuspension technologies for delivery of drugs. Nanosci Nanotech Res 2017; 4(2): 5966. Kilor V, Sapkal N, Daud A, Humne S, Gupta T. Development of stable nanosuspension loaded oral films of glimepiride with improved bioavailability. Int J Appl Pharm 2017; 9(2): 28-33. He J, Han Y, Xu G, Yin L, Neubi MN, Zhou J, et al. Preparation and evaluation of celecoxib nanosuspensions for bioavailability enhancement. RSC Adv 2017; 7: 13053-13064. 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Development and characterization of nanosuspensions of olmesartan medoxomil for bioavailability enhancement. J Pharm Bioall Sci 2011; 3(3): 426-434. Mohd-Fuat AR, Kofi EA, Allan GG. Mutagenic and cytotoxic properties of three herbal plants from Southeast Asia. Trop Biomed 2007; 24(2): 4959. Ravichandran R. Studies on dissolution behaviour of nanoparticulate curcumin formulation. Adv Nanoparticles 2013; 2(1): 51-59. Hussain N, Jaitley V, Florence AT. Recent Advances in the understanding of uptake of microparticulates across the gastrointestinal lymphatics. Adv Drug Deliv Rev 2001; 50(1-2): 107-142. Yuan H, Chen J, Du YZ, Hu FQ, Zeng S, Zhao HL. Studies on oral absorption of stearic acid sln by a novel fluorometric method. Colloids Surf B Biointerfaces 2007; 58(2): 157-164. Gursoy RN, Benita S. Self-emulsifying drug delivery systems (sedds) for improved oral delivery of lipophilic drugs. Biomed Pharmacother 2004; 58(3): 173-182. Liu D, Pan H, He F, Wang X, Li J, Yang X, et al. 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Quality study of pharmaceutical excipient polysorbate 80 / 中国药科大学学报

@#To improve the current quality standards of polysorbate 80 and provide reference recommendations for the revision of the quality standards of polysorbate 80 in the fourth part of China Pharmacopoeia(2015 Edition). A total of 16 batches of polysorbate 80 samples from 6 domestic and foreign production companies were studied, optimize the detection method for some impurities. Adjust the split ratio of the ethylene oxide and dioxane check items, and appropriately increase the concentration of the reference solution solution and change the solvent. In the ethylene glycol and diethylene glycol items, the concentration of the reference solution and the internal standard solution were appropriately increased. The infrared identification and the triethylene glycol check items were added to the quality standards, and we carry out corresponding methodological investigation on the improved method. The results showed that the improved methods had good specificity, precision, linearity and recovery rate. The improved quality standard is more suitable for the detection of polysorbate 80, and can increase the quality standards of polysorbate 80 from safety and standardization.

Nanosuspension enhances dissolution rate and oral bioavailability of Terminalia arjuna bark extract in vivo and in vitro

Objective: To enhance the dissolution rate and oral bioavailability of Terminalia arjuna bark extract by formulating its nanosuspension. Methods: Nanoprecipitation approach was used for the formulation of nanosuspension using polysorbate-80 as a stabilizer. The formulated nanosuspension was assessed for particle size, polydispersity index, zeta potential value and for in vitro dissolution study. Oral bioavailability studies were carried out in Wistar male albino rats by administering a single dose (50 mg/kg. b. wt) of the formulated nanosuspension and coarse suspension. The storage stability of the formulated nanosuspension was determined after three months of storage at room temperature and under the refrigerated condition. Mutagenicity assay was carried out to evaluate the toxicity of the formulated nanosuspension using two mutant strains (Salmonella typhimurium TA100 and Salmonella typhimurium TA98).Results: The mean particle size of the formulated nanosuspension was 90.53 nm with polydispersity index and zeta potential values of 0.175 and ?15.7 mV, respectively. Terminalia arjuna nanosuspension showed improved dissolution rate and 1.33-fold higher oral bioavailability than its coarse suspension. The formulated nanosuspension also showed better stability under the refrigerated condition and was non-mutagenic against both strains. Conclusions: Our study demonstrates that nanosuspension technology can effectively enhance the dissolution rate and oral bioavailability of Terminalia arjuna bark extract.

Analysis on Safety of Polysorbate 80 as a Solubilizing Excipient in Traditional Chinese Medicine Injections / 中国实验方剂学杂志

There are safety problems in traditional Chinese medicine(TCM) injections. Most of the adverse reactions of TCM injections are very similar to those of hypersensitivity reactions. The hypersensitivity reaction of macromolecular substances in TCM injections has also been confirmed by experiments. Polysorbate 80 (trade name of tween-80) is a solubilizing excipient commonly used in TCM injections. Polysorbate 80 contains macromolecular impurities and the material basic research is not clear enough, and its quality problems need to be solved urgently. Based on this, the authors concluded that the macromolecular impurities in polysorbate 80 were the important material basis for the safety of TCM injections contained polysorbate 80. In this paper, the research progress of application of polysorbate 80 as a solubilizing excipient in TCM injections was reviewed and analyzed, so as to provide ideas for improving the safety of TCM injections contained polysorbate 80 and to promote the healthy development of TCM injections.

Research progress on quality evaluation and assay methods of polysorbate 80 / 药物评价研究

Research progress ofpolysorbate 80 was summarized,including standards,chemical composition and impurity test.Assay methods of polysorbate 80 in preparations were also summarized.In addition,characteristics of the current technologies were discussed.Compositions of polysorbate 80 could be detected directly by spectrophotometry,SEC-ELSD or LC-MS methods.In addition,they could be analyzed indirectly by determing the hydrolysates with HPLC-UV or GC methods.In recent years,remarkable progress has been achieved in chemical composition determination and impurity test ofpolysorbate 80.But assay method of the excipient in related drugs need further study,due to large difference between purities ofpolysorbate 80 raw materials.

Research progress on quality evaluation and assay methods of polysorbate 80 / 药物评价研究

Research progress ofpolysorbate 80 was summarized,including standards,chemical composition and impurity test.Assay methods of polysorbate 80 in preparations were also summarized.In addition,characteristics of the current technologies were discussed.Compositions of polysorbate 80 could be detected directly by spectrophotometry,SEC-ELSD or LC-MS methods.In addition,they could be analyzed indirectly by determing the hydrolysates with HPLC-UV or GC methods.In recent years,remarkable progress has been achieved in chemical composition determination and impurity test ofpolysorbate 80.But assay method of the excipient in related drugs need further study,due to large difference between purities ofpolysorbate 80 raw materials.

Validation of Microbial Limit Test for Compound Benzocaine Gel / 中国药师

Objective: To establish a microbial limit test method for compound benzocaine gel.Methods: According to the general principles of Chinese Pharmacopoeia (2015 edition), method applicable experiments were performed respectively for the routine method, neutralization method and dilution & neutralization method.Using the recovery ratio of test bacteria as the index, the medium of dilution & neutralization method was adopted in the total aerobic microbial count and total combined yeasts and molds count, and the neutralization method was used for the control bacteria detection.The neutralizing agents were polysorbate 80 and lecithin.Results: The method of dilution & neutralization could eliminate the bacteriostasis of the drug, and the recovery ratio of each test bacteria was within the range of 0.5-2.0, which was in line with the requirements of Chinese Pharmacopoeia.Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus could be detected out in the positive control bacterial test,and bacteria were not detected out in the negative test.Conclusion: The method can be used for the microbial limit test for compound benzocaine gel.

Neutralization of Lecithin and Polysorbate-80 in Antibacterial Activity of Traditional Chinese Medicine Preparations / 中国药师

Objective:To investigate the neutralization of lecithin and polysorbate-80 in antibacterial activity of oral solid tradition-al Chinese medicine preparations without Chinese medicinal herb powder. Methods:According to the requirements in Chinese Pharma-copeia (2015 edition), the applicability of microbial counting method was tested in the diluent containing lecithin and polysorbate-80 and the conventional diluent, respectively, and the results were compared. Results: The recovery results of applicability of microbial counting method using neutralizers (lecithin and polysorbate-80) conformed to the standard of Chinese pharmacopoeia (2015 edition). Conclusion:The combination of lecithin and polysorbate-80 has notable neutralization in antibacterial activity of oral solid traditional Chinese medicine preparations without Chinese medicinal herb powder.

Relationship between the chemical composition of polysorbate 80 and its solubilization effect / 中国药学杂志

OBJECTIVE: To determine the solubilization effects of the eight types of chemical components of polysorbate 80 to elucidate its material basis for solubilization, and provide a theoretical basis for the rational use of polysorbate 80. METHODS: The shaking flask method and HPLC method were used to determine the concentrations of four types of water-insoluble drugs in the aqueous solution of the eight types of chemical components of polysorbate 80. RESULTS: The solubilization effects of the eight types of chemical components of polysorbate 80 had significant difference. The polysorbate 80 with the best solubilization effect for different types of water-insoluble drugs differed in their chemical compositions. CONCLUSION: The contents of the chemical components of commercially available polysorbate 80 vary significantly, which is considered to be the material basis for the differences in solubilization effect. The results of this study are valuable for the effective use and quality control of polysorbate 80.

Distribution in mouse tissues of P-80 modified neurotoxin nanoparticles after intranasal administration in rats / 中草药

Objective: To prepare neurotoxin nanoparticles (NT-NP) labeled by FITC and modified by polysorbate-80 (P-80), and to compare the in vivo tissue distribution after intranasal administration in rat. Methods: The FITC labeled P-80-NT-NP and NT-NP were prepared, blood samples were collected after intranasal administration in rats at 5, 15, 30, 60, 120, and 240 min, and the brain, heart, liver, spleen, lung, kidney tissue samples were collected. Using FITC-NT as the index component and fluorescence analysis method, the differences of the nanoparticles distribution in rats tissues after modified by P-80 were investigated. Results: After 5, 15, 30, 60, 120, and 240 min of intranasal administration of P-80-NT-NP and NT-NP, the drug was distributed in plasma, heart, liver, spleen, lung, kidney, and brain. After 120 min of administration, the NT concentration in plasma and brain of P-80-NT-NP group was higher than that of NT-NP group, while the NT concentration in liver was lower than that in NT-NP group, with significant difference. Conclusion: P-80 can effectively increase the nasal absorption of NT-NP into the brain and other tissues, which provides the basis for further development and application of preparation of the drug targeting to brain.

Study of degranulation induced by the chemical composition of polysorbate 80 in a rat mast (RBL-2H3) cell line / 中国药学杂志

OBJECTIVE: To study directive effects of eight types of the chemical composition of polysorbate 80 on the degranulation in the RBL-2H3 rat mast cell line and to illuminate the material basis for its pseudoallergy. METHODS: RBL-2H3 cells were cultured and treated with varying dosage of eight types of the chemical composition of polysorbate 80. And then the amount of the released β-hexosaminidase was detected. MTT assay was used to determine the cytotoxicity of eight types of the chemical composition of polysorbate 80 which causing RBL-2H3 cells degranulation. RESULTS: PSM, PSD and PIM induced RBL-2H3 cells' degranulation in a concentration-dependent manner. Degranulation was not detected in cells treated with PS and PI. PSTri, PSTetra and PSD weakly induced the degranulation of RBL-2H3 cells in a concentration-dependent manner. PSM, PSD and PIM showed cytotoxicity with concentration-dependent manner, while PS and PI had no cytotoxicity. PSTri, PSTetra and PSD showed low toxicity cytotoxicity in a concentration-dependent manner. CONCLUSION: PIM is a major source of toxic substances polysorbate 80, the limit of which must be controlled.

Investigation on determination of the content of polysorbate 80 in traditional Chinese medicine injections / 中国药学杂志

OBJECTIVE: To point out the problems of the SEC-ELSD method for determination of polysorbate 80 in traditional Chinese medicine injections, and discuss the method. METHODS: The retention behaviors of eight types of chemical compositions in polysorbate 80 in size exclusion chromatography were studied according to the chromatographic conditions of SEC-ELSD method. RESULTS: Not all of the chemical compositions had same chromatographic retention time, some of them showed no chromatographic peaks in the chromatographic conditions of SEC-ELSD method. CONCLUSION: In order to ensure the rational use of SEC-ELSD method, researchers must be concerned about the state of the surfactant molecules in the mobile phase and the interaction between surfactant molecules with the stationary phase. A new standard of polysorbate 80 must be implemented, the core of which is controlling the chemical compositions.

Application of Polysorbate 80 in Microbial Limit Test / 中国药师

Objective:To provide appropriate test liquid preparation methods for antibacterial or water-insoluble drugs, and estab-lish a convenient and effective method for microbial limit test. Methods:Using function of polysorbate 80 as neutralization, emulsifica-tion, solubilization and microbial protection, We carried out the liquid preparations were prepared and microbial limit test validation was carried out for 11 kinds of hospital drugs. Results:The bacteria recovery rate and the control bacteria test were in accordance with the requirement of pharmacopoeia. Conclusion:Polysorbate 80 can be used in the microbial limit test of the drugs.

Preparation and purity determination of the reference standards for chemical compositions of polysorbate 80 / 中国药学杂志

OBJECTIVE: To establish a new method for the preparation and purity determination of the reference standards for eight types of chemical compositions in polysorbate 80. METHODS: Polyoxyethylene 24 sorbitan oleate and polyoxyethylene 12 isosorbide oleate were synthesized respectively. Medium and low pressure preparative chromatography and flash column were used to separate and purify the constituents, and their structures were elucidated by IR, Raman, MS, 1H-NMR and 13C-NMR. RESULTS: Eight single chemical compositions of polysorbate 80 were obtained, the purities of which were greater than 99.5% tested by HPLC, meeting the requirement as a reference standard. CONCLUSION: A directed synthesis together with a medium and low pressure preparative chromatography method is established for the first time which can accurately and rapidly prepare the reference standards for eight chemical compositions of polysorbate 80. And for the first time the spectral data of the eight chemical components were reported.

Analysis of the chemical composition of polysorbate 80 / 中国药学杂志

OBJECTIVE: To establish a method for the separation and identification of the components in polysorbate 80 and to lay the foundation for identifing the sensitinogen of polysorbate 80. METHODS: Polysorbate 80 samples from both domestic and foreign manufacturers were collected. The separation condition was first established using HPLC-ELSD, and then the chemical composition of each chromatographic peak was identified by HPLC-MS. RESULTS: The chemical components with different types of structures were efficiently separated, and each chromatographic peak was properly assigned. Polysorbate 80 samples of different batches and manufacturers showed different chemical compositions and relative contents. There was illegal blending during the production of polysorbate 80. CONCLUSION: The proposed method is rapid and simple, which is not only suitable for the process control of polysorbate 80 but also lays foundation for further investigation of the physical and chemical properties of the components thus to find the sensitinogen. Copyright 2012 by the Chinese Pharmaceutical Association.