ABSTRACT
Foot and mouth disease is a highly contagious viral disease of cloven-hoofed animals that has a significant economic impact on livestock. A recent outbreak was detected and recorded as exotic strain of foot and mouth disease virus SAT2 (Serotype SAT2, topotype VII, Lib-12 lineage). The emergency vaccine was produced and assessed in vivo and large number of vaccine batches were urgently needed. The present work was aimed to provide a rapid evaluation of inactivated foot and mouth disease SAT2 oily vaccine to exclude the unsatisfactory batches during emergency circumstances and to reduce time, effort and cost. The extraction of foot and mouth disease antigen content from oily adjuvanted vaccine was carried out using isopropyl myristate and benzyl alcohol methods. The extracted viral antigen was identified by foot and mouse disease serotyping ELISA and 146S content was quantified using sucrose density gradient analysis. Evaluations were carried out instantly and at 2h, 6h and 24h. The results indicated the efficiency of benzyl alcohol to breakdown the oil emulsion either MONTANIDE™ ISA 206 VG or MONTANIDE™ ISA 50 V2, while the isopropyl myristate was efficient for MONTANIDE™ ISA 50 V2 only. The identification and quantification of 146S for extracted antigen using benzyl alcohol indicated significant stable records at different time intervals for the vaccine batches, while the extraction using isopropyl myristate indicated unstable records at different time intervals. It was concluded that the evaluation of monovalent foot and mouse disease vaccine could be conducted in vitro, using serotyping ELISA and quantification of 146S for the extracted antigen, either using benzyl alcohol or isopropyl myristate (MONTANIDE™ ISA50 V2 only), with the consideration that 146S content should not less than 4 μg/mL(AU)
La fiebre aftosa es una enfermedad viral altamente contagiosa de los animales de pezuña hendida que tiene un impacto económico significativo en el ganado. Se detectó un brote reciente que se registró como causado por una cepa exótica del virus de la fiebre aftosa (serotipo SAT2, topotipo VII, linaje Lib-12). La vacuna de emergencia se elaboró y evaluó in vivo, existiendo una urgente necesidad de contar con un gran número de lotes de la misma. El presente trabajo tuvo como objetivo proporcionar una evaluación rápida de la vacuna oleosa inactivada (SAT2) contra la fiebre aftosa, para excluir los lotes insatisfactorios durante circunstancias de emergencia, reduciendo tiempo, esfuerzo y costo. La extracción del contenido de antígeno de fiebre aftosa, de la vacuna oleosa adyuvada, se llevó a cabo utilizando miristato de isopropilo y alcohol bencílico. El antígeno viral extraído se identificó utilizando un ELISA de serotipificación y se cuantificó el contenido de 146S mediante análisis de gradiente de densidad de sacarosa. Las evaluaciones se realizaron de forma instantánea y a las 2h, 6h y 24h. Los resultados indicaron la eficacia del alcohol bencílico para separar la emulsión de aceite para MONTANIDE ™ ISA 206 VG o MONTANIDE™ ISA 50 V2, mientras que el miristato de isopropilo fue eficaz para MONTANIDE™ ISA 50 V2 únicamente(AU)
Subject(s)
Animals , Enzyme-Linked Immunosorbent Assay/methods , Foot-and-Mouth Disease , Vaccines , EgyptABSTRACT
Objective: To establish an HPLC method to determine the content of benzyl alcohol in metamizole sodium injection. Methods: The determination was performed on an Xtimate C18(250 mm×4. 6 mm,5 μm)column. Phosphate buffer (dissolving 6. 0 g sodium dihydrogen phosphate in 1000 ml water, adding 1ml triethylamine, adjusting pH to 7. 0 with sodium hydroxide solution) -methanol (75: 25) was used as the mobile phase at a flow rate of 1. 0 ml·min-1. The detection wavelength was 254 nm. The column tamprture was 30 ℃ and the sample size was 5 μl. Results: The linear range of benzyl alcohol was 76. 88-269. 08 μg·ml-1( r=0. 998 5)with the average recovery of 98. 77% (RSD=0. 77% ,n=9). Conclusion: The method is accurate and reproducible for the content determination of benzyl alcohol in metamizole sodium injection.
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OBJECTIVE: To establish an HPLC method for determination of the preservatives in betamethasone injectable suspension, including benzyl alcohol, methylparaben and propylparaben. METHODS: The determination method was developed with 5 mmol·L-1 ammonium acetate(containing 1% triethylamine and adjusted to pH 5.0 with glacial acetic acid) as mobile phase A and acetonitrile as mobile phase B, and gradient elution was performed at a flow rate of 1.0 mL·min-1. The column temperature was maintained at 30℃ and the detection wavelength was set at 257 nm. RESULTS: The resolutions between the three main peaks and adjacent peaks were greater than 1.5. The calibration curves of benzyl alcohol, methylparaben and propylparaben showed good linearity in the ranges of 0.175 9-1.758 6 mg·mL-1(r=1.000 0), 0.022 3-0.223 1 mg·mL-1(r=0.999 8), 0.004 35-0.043 47 mg·mL-1(r=1.000 0), respectively. The average recovery rates were 98.98%, 99.38% and 100.07%, and the RSDs were 0.6%, 1.3% and 1.0%(n=9), respectively. The results of test of influencing factors showed that the contents of hydroxyphenylmethyl ester and propylparaben decreased at high temperature. CONCLUSION: The developed method is proved by validation to be applicable for the quality control of the preservatives in betamethasone injectable suspension.
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Objective:To determine the related substances in benzalkonium chloride used as a pharmaceutical adjuvant, and com-pare the quality at home and abroad. Methods:An HPLC method was used with an ODS-HYPERSIL C18 column(250 mm × 4. 6 mm, 5 μm). The detection wavelength was 210 nm and 257 nm. The flow rate was 1. 0 ml· min-1 and the column temperature was 30℃. The injection volume was 20 ml. The mobile phase was A ( dissolving 1. 09 g sodium1-hexanesulfonate and 6. 9 g sodium dihydrogen phosphate in water, adjusting pH to 3. 5 with phosphoric acid and diluting to 1 000. 0 ml) and B ( methanol) with gradient elution. Results:The content of benzaldehyde in the samples at home and abroad was low. The content of benzyl alcohol in the samples from a-broad was qualified, which was significantly higher than that in the domestic samples. The content of benzyl chloride in the domestic samples was higher than that in the samples from abroad. Conclusion:The method is simple and fast, which is suitable for comparing the related substances of domastic and imported samples. At the same time, the study provides basis for enterprises to choose benzalko-nium chloride rationally.
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OBJECTIVE:To establish the quality standard for Gastrodia elata hency-fired tablet. METHODS:TLC was used to identify the gastrodin and benzyl alcohol and determine the content of moisture,ash and extract;HPLC was used to determine the content of gastrodin and benzyl alcohol. Column was Diamonsil C18 with mobile phase of acetonitrile-0.05% phosphate(3:97,V/V) at flow rate of 1.0 ml/min,detection wavelength was 220 nm,column temperature was 25 ℃ and volume injection was 10 μl. RE-SULTS:The TLC of gastrodin and benzyl alcohol showed clear spots and good separation. The moisture content40.0%. Linear range of gastrodin was 25.2-126.0,12.7-63.5 μg/ml(r=0.999 9),respectively;RSDs of precision,reproducibility and stability tests were lower than 2.0%;recovery was 99.49%-102.40%(RSD=1.09%,n=6), 98.75-102.63%(RSD=1.53,n=6),respectively. CONCLUSIONS:The method is simple and good reproducibility,and can be used for the quality control of Gastrodia elata hency-fired tablet.
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To establish a GC method to determine the related substances in pharmaceutical excipient benzyl alcohol. Methods:A GC method was used with an Agilent DB-wax eapillary column(0. 32 mm × 30 m,1. 8 μm)and programming temperature. The initial temperature was 50℃, and then raised to 220℃ with a rate of 5℃·min-1 and maintained for 35min. The detector was FID. The temperature of the injection port was 200℃,and the detector temperature was 310℃. The results were confirmed by GC-MS. Results:Within a certain range,the peak area and concentration of every impurity had a good linear relationship (r≥0. 999 9). The recovery was between 96. 1% and 102. 7%. The quantitative limit was between 1. 37-3. 63 ng. Toluene, benzyl chloride, benzalde-hyde and benzyl ether were found out in the samples. Conclusion:The method is accurate and convenient, and suitable for the quanti-tative determination of related substances in pharmaceutical excipient benzyl alcohol.
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OBJECTIVE: To study the chemical constituents of Dendrobium officinale protocorm. METHODS: The compounds were isolated and purified by means of column chromatography, and their structures were identified on the basis of spectroscopic data and comparison with standard literature data. RESULTS: Eight compounds were isolated from the(CH3)2CO, CHCl3 and 95% EtOH fractions of the EtOH extract, and identified as 1-O-p-feruloyl-β-D-glucopyranoside(I), arillatose B(II),4-β-D-glucopyranosy-loxy)benzyl alcohol(III), 4-hydroxymethyl-2,6-dimethoxyphenyl-β-D-glucopyranoside(IV), n-hexatriacontanoic acid(V), n-heptaco-sanol(VI), β-sitosterol(VII), and cycloart-23-ene-3β,25-diol(VIII). CONCLUSION: All these compounds were isolated from the protocorms of D. officinale for the first time. Compounds I, II, III, IV, and VIII were obtained from Dendrobium genus for the first time.
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OBJECTIVE:To establish HPLC method for the content determination of bacteriostatic agent benzyl alcohol in Clindamycin phosphate injection.METHODS:The determination was performed on CAPCELL PAK C8 column using potassium dihydrogen phosphate solution-acetonitrile(775:225)as mobile phase at flow rate of 1.0 mL?min-1.The detection wavelength was set at 210 nm.RESULT:The linear range of benzyl alcohol was 4.175～41.75 ?g?mL-1(r=0.999 9)with an average recovery of 100.5%(n=9),RSD=0.09%.CONCLUSION:The method is accurate and reproducible for the content determination of benzyl alcohol in Clindamycin phosphate injection.
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[Objective]To explore the mechanism of hip joint capsule contracture induced by intragluteal injection. [Method]Thirty young Chinese rabbits were divided into four groups of A(n=6),B(n=6),C(n=6) and D(n=12) at random.Rabbits in groups A,B and C were treated with deep intragluteal injection of penicillin dissolved in 2% benzyl alcohol(group A),2% benzyl alcohol(group B)and penicillin dissolved in physiologic saline(group C).Rabbits in group D were treated with immobilization of unilateral hip joint and deep intramuscular injection of physiologic saline on the homolateral buttock.Each rabbit was injected twice a day for 20 days.Specimens were obtained at 30,60 and 90 days after first injection and the evaluation of the functions of hind limbs was obtained at the same time.The thickness of hip joint capsules were measured with micrometer and sections of joint capsules were stained by Sirius red and were observed with microscope and polarization microscope.[Result]Capsules of hip joint in group A and B became thicker and contractural,and the change of joint capsules was reflected in the change of motion of hind limbs.Sections in groups A and B showed shrinked capsules with thick collagens in disordered arrangement;while there were no gluteus contracture and contracture of joint capsule in the other groups.[Conclusion]Deep intragluteal injection of penicillin dissolved in 2% benzyl alcohol or 2% benzyl alcohol into young rabbits can cause contracture of hip joints.The contracture of joint capsule is largely induced by tissue trauma and repair of joint capsules.The limited locomotion of hip joint induced by the contracture of gluteal muscle does not play an important role in the contracture of joint capsule.