ABSTRACT
Objective: To investigate the effects and mechanism of Zhenzhu Mingmu Eye Drops (ZMED) on human corneal epithelial cells (HCEC) injury model induced by high permeability. Methods: The dye eyes disease model was established using 110 mmol/L NaCl to stimulate HCEC, and different concentration of ZMED was given to the treatment group. MTT assay and lactate dehydrogenase (LDH) leakage were used to evaluate the protection effects of ZMED on HCEC. The effects of ZMED on apoptosis of HCEC were analyzed by flow cytometry. AO/EB double staining and Hoechst 33258 analysis were used as sensitive assays for apoptosis. Moreover, HCEC treated with ZMED were subjected to Western blotting for protein levels of Caspase-9, Caspase-3 and PARP. Results: ZMED significantly increased cell viability of HCEC and decreased the LDH release in a dose-dependent manner. ZMED dramatically decreased the apoptosis rate. Furthermore, ZMED down-regulated the levels of cleaved Caspase-9, cleaved Caspase-3 and cleaved PARP. Conclusion: ZMED could protect HCEC against hypermeability, which may be related to the inhibition of Caspase-9/Casepase-3 signaling pathways.
ABSTRACT
Objective: Analysis for the differences of protein polypeptide ingredients in Zhenzhu Mingmu Eye Drops and Zhenzhuye in order to provide the reference data to ensure the stability of production process of crude material and effectively control the quality of Zhenzhu Mingmu Eye Drops. Methods: Polypeptide ingredients in Zhenzhu Mingmu Eye Drops and Zhenzhuye were identified by UPLC-Q-TOF MS in positive mode using an Acquity UPLC BEH C18 (50 mm×2.1 mm, 1.7 μm) column. Mobile phase is acetonitrile-0.1% methanol; Contents of 16 amino acids were detected by pre-column derivatization RP-HPLC method. Results: Molecular weight (MW) of polypeptide and the contents of the amino acids in Zhenzhu Mingmu Eye Drops and Zhenzhuye were significantly different. In the five manufacturers, MW of polypeptide was less than 2000 in Zhenzhuye samples from the three manufacturers, while MW of polypeptides were 500-4746 for samples of the other two manufacturers and MW of main polypeptides were higher than 2000.MW ranges of main polypeptides in Zhenzhu Mingmu Eye Drops from two manufacturers were 2000-4000 and beyond 4000. There were not the polypeptides with MW beyond 4000 in samples of the other five manufacturers. The content percentages of 16 amino acids in Zhenzhu Mingmu Eye Drops from the seven manufacturers were obviously different. The mass concentration ratios of proline and alanine in Zhenzhuye produced by acid/enzyme hydrolysis from two manufacturers are beyond 1.For the other five manufacturers with water extract technique, the mass concentration ratios of proline and alanine are below 1. Conclusion: The quality standard of Zhenzhuye should be revised and water extract should be applied in preparation process to ensure the effective ingredients polypeptides (500-5000). In quality standard of Zhenzhu Mingmu Eye Drops and Zhenzhuye, the mass concentration ratio of proline/alanine and the total contents of amino acids should be added as check and content determination items, respectively, so as to effectively control the quality of Zhenzhu Mingmu Eye Drops and ensure its validity.
ABSTRACT
Objective:To evaluate the antimicrobial effectiveness of Zhenzhu Mingmu eye drops according to the requirements de-scribed in Chinese pharmacopoeia (2010 edition). Methods:Totally 7 batches of Zhenzhu Mingmu eye drops from 7 enterprises were evaluated. All the samples were tested according to the requirements described in Chinese pharmacopoeia, and the testing points of 6h and 24h were added. Results:As for bacteria, the reduction value of staphylococcus aureus in 7 days treated with Zhenzhu Mingmu eye drops from enterprise e was 0. 61g, and that in 14 days was 1. 01g. That in 7 days treated with the eye drops from the other enter-prises was all above 1. 01g, that in 14 days was above 3. 01g, and that in 14-28 days kept stable. As for fungi, the number was stable from the beginning with the treatment of the eye drops from the seven enterprises. Conclusion:Although the type and content of bacte-riostatic agent in domastic Zhenzhu Mingmu eye drops are the same, there is difference in the antimicrobial effectiveness of the product from different enterprise. The eye drops from one enterprise can't meet the requirements for type I product described in Chinese pharma-copoeia,and the others meet the requirements.