The cytotoxic effects of latanoprost, travoprost, and bimatoprost were dependent on the BAC concentration in their formulations. BACmix was cytotoxic at the concentrations above those corresponding to 0.001% BAC in ophthalmic medications. PF tafluprost was the least toxic of the drugs tested. Within studied BAC homologs, those with longer alkyl chain and higher lipophility penetrated effectively into rabbit external ocular tissues.

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Synthetic biodegradable polymers have many potential therapeutic applications. In ophthalmology, biodegradable polymers have been used as viscoelastic agents and surgical implants. Other potential applications include controlled release of drugs and growth factors, gene therapy, and tissue engineering. In the present study, in vitro biocompatibility of three biodegradable polymers, 50:50 PDLGA, 85:15 PDLGA, and Inion GTR membrane was evaluated in comparison to tissue culture polystyrene by investigating cell proliferation and potential acute toxicity by the WST-1 cytotoxicity/cell proliferation test, the ATP test, and the lactate dehydrogenase (LDH) test. Evaluations were conducted with cell line cultures from various ocular tissues, human corneal epithelial cells (HCE), rabbit stromal fibroblasts (SIRC), bovine corneal endothelial cells (BCE), human conjunctival epithelial cells (IOBA-NHC), and human retinal pigment epithelial cells (ARPE-19) by direct contact studies by plating the cells on the polymer film specimens in 96-wells. The proliferation results show that cell lines from various ocular tissues attached and grew on PDLGA 50:50, PDLGA 85:15, and Inion GTR membrane. Cytotoxicity experiments with the LDH and ATP tests showed no or extremely slight toxic adverse effects. These polymers have potential to be used as scaffolds in cell transplantation devices or as surgical implants.

Lipase biocatatysis was investigated as a tool for the production of butyl oleate and rapeseed oil 2-ethy]-I -hexy[ ester by esterification and transesterification, respectively. We screened 25 commercially available [ipases and found that butyl oleate was produced at high yields from oleic acid and 1butanol by [ipases from Candida rugosa, Chromobacterium viscosum, Rhizomucor miehei, and Pseudomonas fluorescens. The initial water content of the system, lipase quantity, and the molar ratio of t-butanol to oleic acid were important factors in influencing the ester yield. In general, no ester was formed without the addition of water. The exception was Ch. viscosum lipase, which yielded 98% of ester in 12 h with t-butanol excess without additional water. The addition of 3.2% water increased the initial rate of reaction. With an oleic acid excess and only 0.3% lipase, C. rugosa and R. miehei lipases yielded 94 and 100% esters with initiaf water contents of 3.2 and 14%, respectively. Lipase-catatyzed alcoholysis of [ow-erucic acid rapeseed oil and 2-ethyl-I -hexanol without additional organic solvent also was studied in stirred batch reactors. In this case, C. rugosa lipase was the best biocatalyst with an optimal 2-ethyl-1 -hexanol to rapeseed oil molar ratio of 2.8, a minimum of 1.0% added water, and 37°C. An increase in temperature up to 55°C increased the rate of reaction but did not affect the final ester yield. The enzyme was inactivated at 60°C. Under optimal conditions, the ester yield increased from 88% in 7 h to nearly complete conversion in 1 h when the lipase content was increased from 0.3 to 14.6%. In a 2-kg small pilot scale, up to 90% conversion (97% of theoretical) was obtained in 8 h at 37°C with 3.4% lipase in the presence of Amberlite XAD-7 resin with 3% added water.

Lipase-catalyzed transesterification (alcoholysis) of lowerucic acid rapeseed oil and 2-ethyl-l-hexanol without an additional organic solvent was studied in stirred batch reactors. Of a number of commercially available enzymes investigated, the best results were obtained with a Candida rugosa Upase. The optimal transesterification conditions were an oil/alcohol molar ratio of 1:2.8, a minimum of 1.0% (w/w) added water, and with a temperature of 37-55~ Under the optimal conditions, a nearly complete conversion was obtained in one hour with 14.6% (w/w) lipase, whereas 0.3% (w/w) lipase required 10 h for similar results. The enzyme was inactivated at 60~ KEY WORDS: Alcoholysis, biocatalysis, enzyme, 2-ethyl-l-hexanol, 2-ethyl-l-hexyl ester, lipase, rapeseed oil, transesterification.

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The cytotoxicity of benzalkonium chloride (BAC) and disodium edetate (EDTA) was evaluated in vitro in rabbit corneal epithelial primary cells and in the immortalized human corneal epithelial cell line SV40. Cell injury was assessed by lactate dehydrogenase (LDH) leakage and by reduction of the tetrazolium salt WST-1 to formazan by mitochondrial metabolic activity. Cell cultures were exposed to test compounds both in serum-free and in serum-containing medium. Although WST-1 and LDH tests measured different physiological endpoints, they yielded comparable results. However, the LDH test seemed less reliable due to great variation. The use of serum was found to result in lower toxicity of the compounds in both tests. The rabbit primary cell culture and the human corneal cell line were quite similar in their responses to BAC and EDTA. The human cell line is a promising in vitro alternative in oculotoxicity testing.