Notes

1. Analysis of the mixture composition can be performed by taking samples at intervals. After a 20-fold dilution in water, filter to eliminate the enzyme and analyze by high-performance liquid chromatography (HPLC) on a reverse-phase column (Nucleosil C18, 6 ^m, 250 mm x 4 mm inside diameter; I.C.S., France). Eluent: 50/50 methanol/water; flow rate: 0.5 mL/min; temperature: 33°C; injection volume: 10 ^L; detection: refractometer (RI-Detector 8110, I.C.S.).

2. The yield is defined as the molar ratio of a-butylglucoside lactate formed to initial a-butylglucoside.

3. Butyl-lactate (boiling point [bp] = 186°C) and hexane (bp = 69°C) can be recovered from the organic phase by distillation.

4. Residual a-butylglucoside, butanol, and butyl-lactate are quantified by HPLC (see Note 1). The final L-lactic acid concentration is assayed using the L-lactate kit from Boehringer Mannheim (Germany).

5. Analysis of the mixture composition can be performed by taking samples at intervals. After a 10-fold dilution in methanol (for acid analysis) or in 5 mM sulfuric acid (for a-butylglucoside analysis), the sample is filtered to eliminate the enzyme and analyzed. Linoleic acid, oleic acid, and linolenic acid concentrations are quantified by HPLC chromatography on a reverse-phase column (Nucleosil C18, 6 ^m, 250 mm x 4 mm inside diameter, I.C.S., France). Eluent: 87/13/0.3 methanol/water/trifluoroacetic acid, flow rate: 1.0 mL/min; temperature: 50°C; injection volume: 10 ^L; detection: refractometer. a-Butylglucoside is quantified by HPLC chromatography on an ion-exclusion column (PPH 224 [Brownlee Labs], 4.6 mm inside diameter x 220 mm). Elution conditions are 5 mM sulfuric acid, 0.3 mL/min flow rate, 25°C temperature, 10 ^L injection volume, and a refractometer for detection.

6. The yield is defined as the molar ratio of total a-butylglucoside esters formed to initial a-butylglucoside (or total acids).

7. Exact concentration of each a-butylglucoside ester (linoleate, oleate, and linolenate) can be quantified by HPLC (see Note 5). The content of each a-butylglucoside ester is the same as the content of the corresponding acid in the commercial mixture of unsaturated fatty acids.

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