Enzyme-catalyzed Synthesis of Vitamin E Succinate Using a Chemically Modified Novozym-435

Vitamin E succinate was synthesized in organic solvents using a modified Novozym-435 as catalyst.In order to improve the catalytic performance of Novozym-435,the enzyme was modified using acetic anhydride, propionic anhydride and succinic anhydride separately.We found that both the hydrolytic activity and the thermal stability of the modified Novozym-435 were enhanced compared with the unmodified enzyme.The modified Novozym-435 catalysts were used to synthesize the succinate derivative of vitamin E.Compared with the native Novozym-435,the catalytic activity of the modified novozym-435 in promoting the synthesis of vitamin E succinate was dramatically increased,with the novozym-435 modified with succinic anhydride（N435-S）as the most active catalyst.Conditions for the synthesis of vitamin E succinate were also optimized.A mixture of tert-butanol and DMSO（volume ratio of 2︰3）was the most suitable medium for the reaction,whereas the appropriate molar ratio of vitamin E to succinic anhydride and reaction temperature were 1︰5 and 40°C,respectively.Under these reaction conditions,the yield of vitamin E succinate reached 94.4%.N435-S could be reused for five batches.

Effect of vitamin E succinate on the proliferation of human breast cancer cells

Objective：Tn investigate the growth inhibition and apoptosis induced by vitamin E suceinate （VES） on human breast cancer cells and to analyze the possible mechanism in this process. Methods： Human breast cancer cell line Bcap-37 was treated with VES for 12, 24 and 48 h at the concentrations of 5, 10and 20 μg/ml. Then MTT assay was employed to detect the inhibitory effect of VES on the growth of breast cancer cells. Flow cytometry was used to analyze the cell cycle and apoptosis. To find out whether the Fas/FasL pathway was involved in this process, RT-PCR and flow cytometry assay were used to detect the Fas expression at the mRNA and protein level. Results： VESexhibited a significant inhibitory effect on the growth of human breast cancer cells, presenting in a dose- and time-dependant manner. The apoptotic rate of Bcap 37 cells was 0.6%, rose to 21.0% and 37.5% after treated with VES for 24 and 48 h at the concentration of 20 μg/ml. Fas mRNA transcription was upregulated after VES treatment and cell surface Fas expression increased according to the flow cytometry assay. Concluslon：Significant growth inhibition and apoptosis are induced in human breast cancer cells after treated with VES. The modulation of Fas/FasL pathway may related to the upregulation of Fas molecule on the cancer cell surface.

Synthesis of Vitamin E Succinate from Candida rugosa Lipase in Organic Medium

A screening of commercially available lipases for the synthesis of vitamin E succinate showed that lipase from Candida rugosa presented the highest yield. The synthesis of vitamin E succinate in organic solvents with dif- ferent lgP values ranging from -1.3 to 3.5 was investigated. Of particular interest was that dimethyl sulfoxide （DMSO） with the lowest lgP exhibited the highest yield among all the organic solvents used. It suggests that lgP is incapable of satisfactorily predicting the biocompatibility of organic solvents due to the complexity of enzymatic reaction with hydrophilic and hydrophobic substrates in organic solvent. Effects of different operating conditions, such as molar ratio of substrate, enzyme concentration, reaction temperature, mass transfer, and reaction time were also studied. Under the optimum conditions of 10 g/L enzyme, a stirring rate of 100 r/min, a substrate molar ratio of 5：1 at 55℃ for 18 h, a satisfactory yield（46.95%） was obtained. The developed method has a potential to be used for efficient enzymatic production of vitamin E succinate.

Efficient drug delivery and anticancer effect of micelles based on vitamin E succinate and chitosan derivatives

Nanocarriers have emerged as a promising cancer drug delivery strategy.Multi-drug resistance caused by overexpression of multiple-drug excretion transporters in tumor cells is the major obstacle to successful chemotherapy.Vitamin E derivatives have many essential functions for drug delivery applications,such as biological components that are hydrophobic,stable,water-soluble enhancing compounds,and anticancer activity.In addition,vitamin E derivatives are also effective mitocan which can overcome multi-drug resistance by binding to P glycoproteins.Here,we developed a carboxymethyl chitosan/vitamin E succinate nano-micellar system(O-CMCTS-VES).The synthesized polymers were characterized by Fourier Transform IR,and 1H NMR spectra.The mean sizes of O-CMCTS-VES and DOX-loaded nanoparticles were around 177 nm and 208 nm.The drug loading contents were 6.1%,13.0%and 10.6%with the weight ratio of DOX to O-CMCTS-VES corresponding 1:10,2:10 and 3:10,and the corresponding EEs were 64.3%,74.5%and 39.7%.Cytotoxicity test,hemolysis test and histocompatibility test showed that it had good biocompatibility in vitro and in vivo.Drug release experiments implied good pH sensitivity and sustained-release effect.The DOX/O-CMCTS-VES nanoparticles can be efficiently taken up by HepG2 cancer cells and the tumor inhibition rate is up to 62.57%.In the in vivo study by using H22 cells implanted Balb/C mice,DOX/O-CMCTS-VES reduced the tumor volume and weight efficiently with a TIR of 35.58%.The newly developed polymeric micelles could successfully be utilized as a nanocarrier system for hydrophobic chemotherapeutic agents for the treatment of solid tumors.

Vitamin E Succinic Acid enhances the effect of mDRA-6 to eradicate leukemia cells by inducing apoptosis

Objective: The aim of our study was to detect whether Vitamin E Succinic Acid (VES) could regulate the expression level of DR5 in the cells and further elucidate the potential mechanisms involving that VES enhances the effect of mDRA-6 to eradicate leukemia Raji and K562 cells. Methods: MTT method was used to detect the growth inhibition of VES and mDRA-6 to Raji and K562 cells. Annexin V-FITC/PI double staining assay was used to analysis the apoptosis of leukemia cell. Flow cytometry was used to detect the cell surface DR5 expression. Immunoblotting technique was used to detect the DR5 protein expression. Results: MTT detection showed that 10 μmol/L mDRA-6 on the cell death rates of Raji and K562 cells were 21.98% and 5.23%, respectively. While increasing concentration of VES (5 μmol/L, 10 μmol/L, 20 μmol/L) and mDRA-6 both on the cell viability of Raji or K562 cells, the mortality of Raji cells elevated to 24.67%, 35.65% (P<0.01) and 40.22% (P<0.01), respectively. Similarly, the mortality of K562 cells increase to 6%, 7.89% (P<0.01) and 8.67% (P<0.01), respectively. To further specify the increased cell death rate induced by mDRA-6 and VES, the treated cells were analyzed by Annexin-V/PI staining assay. As shown in Fig. 1, the apoptosis rates of Raji and K562 cells treated with 2 μg/mL mDRA-6 for 12 h were 20.79% and 7.74%. Compared with this, the proportion of apoptotic cells increased upon exposure to 2 μg/mL mDRA-6 combination with 10 μmol/L VES, the apoptosis rates of Raji and K562 cells were 43.18% and 16.99%, respectively. To examine the anticancer effects of a combination strategy based on mDRA-6 and VES. We analyzed whether VES could elevated the expression level of DR5 on Raji and K562 cytomembrane by FACS. Interestingly, after treated with 10 μmol/L VES for 12 h, the expression level of DR5 on Raji and K562 cell surface increased from 50.66% to 70.08%, and 15.02% to 16.38%, respectively. Immune imprinting technology test showed that, different concentrations of VES could increase Raji and K562 cell DR5 protein expression. Conclusion: VES enhances the effect of mDRA-6 to eradicate leukemia Raji and K562 cells. The proper mechanism is VES could enhance the Raji and K562 cell membrane expression of DR5, and VES can also enhance the DR5 protein expression of cells.