Optimization of carbohydrate fatty acid ester synthesis in organic media by a lipase from Candida antarctica.

The effect of solvents and solvent mixtures on the synthesis of myristic acid esters of different carbohydrates with an immobilized lipase from C. antarctica was investigated. The rate of myristyl glucose synthesized by the enzyme was increased from 3.7 to 20.2 micromol min(-1) g(-1) by changing the solvent from pure tert-butanol to a mixture of tert-butanol:pyridine (55:45 v/v), by increasing the temperature from 45 degrees C to 60 degrees C, and by optimizing the relative amounts of glucose, myristic acid, and the enzyme preparation. Addition of more than 2% DMSO to the tert-butanol:pyridine system resulted in a reduction of enzyme activity. Lowering the water content of the enzyme preparation below 0.85% (w/w) resulted in significant decreases in enzyme activity, while increasing the water content up to 2.17% (w/w) did not significantly affect the enzyme activity. The highest yields of myristyl glucose were obtained when an excess of unsolubilized glucose was present in the reaction system. In this case, all of the initially solubilized and a significant amount of the initially unsolubilized glucose was converted to the ester within 24 h of incubation, resulting in a myristyl glucose concentration of 34 mg/mL(-1). Myristic acid esters of fructose (22.3 micromol min(-1) g(-1)), alpha-D-methyl-glucopyranoside (26.9 micromol min(-1) g(-1)) and maltose (1.9 micromol min(-1) g(-1)) could also be prepared using the tert-butanol:pyridine solvent system. No synthesis activity was observed with maltotriose, cellobiose, sucrose, and lactose as substrate.

Two-step enzymatic synthesis of maltooligosaccharide esters.

Glucose and maltose esters were synthesised in organic media by employing a lipase (E.C. 3.1.1.3) from Candida antarctica. In a second reaction step, a transglycosylation catalysed by a cyclodextrin glycosyltransferase (E.C. 2.4.1.19) from either Paenibacillus sp. F8 or Bacillus sp. strain no. 169 (DSM 2518) extended the degree of polymerisation (DP) of the carbohydrate moieties of the carbohydrate esters. The donor substrates used were either a cyclodextrin, a maltooligosaccharide or starch. The highest rate of low DP maltooligosaccharide ester formation was obtained when starch was used as glycosyl donor and caproyl maltose as glycosyl acceptor. The structures of two of the products were identified by 1H and 13C NMR and MALDI-TOF MS as capronate monoesters of maltotriose and maltotetraose, with the ester bond at C-6 of the second glucose unit from the reducing end.

Ca2+ activated myosin-ATPase in cardiac myofibrils of rainbow trout, freshwater turtle, and rat.

The Ca(2+)-activated myosin-ATPase and its dependence on hypoxia were assessed in freshwater turtle, rainbow trout, and in some cases rat. At 20 degrees C and pH 7.3, the maximal ATPase activity was (mean +/- SEM): turtle 0.040 +/- 0.003, trout 0.090 +/- 0.005, and rat 0.12 +/- 0.004 mmol*min-1*g-1 myofibrillar dry weight. The turnover number was about three times lower for turtle than for trout. Trout is typically active at lower temperatures than turtle, and its myosin-ATPase activity was about three times lower at 10 degrees than at 20 degrees C. Addition of 12 mM phosphocreatine showed that the myosin-ATPase activity covered by myofibrillar creatine kinase was 22 +/- 2% for turtle, 14 +/- 2% for trout, and 69 +/- 5% for rat. At pH 6.8 relative to 7.3, the maximal M-ATPase activity was the same, whereas the Ca(2+)-sensitivity decreased, and more so for trout than for turtle. This difference disappeared, when trout myocardium was examined at 10 degrees C. P(i) (15 mM) affected neither maximal activity nor Ca(2+)-sensitivity. ADP, however, reduced maximal myosin-ATPase activity, and more so in trout than in turtle. In conclusion, the "slow"-type myosin, the low sensitivity of acidification and ADP, and the high creatine kinase/myosin-ATPase ratio in turtle relative to trout accord with the well-known ability of turtle myocardium to work during hypoxia. However, the difference in living temperature between turtle and trout obscures the situation (e.g. inclusion of rat data suggests that the creatine kinase/myosin-ATPase ratio is related to temperature.