Alcohols, secondary, oxidation with yeast

Table 40 The primary and secondary deuterium-tritium and hydrogen-tritium KIEs for the oxidation of benzyl alcohol to benzaldehyde with NAD+ and yeast alcohol dehydrogenase at 25°C.a...

As already mentioned, the secondary alcohols that are obtained are optically active. It should be stressed that the reduction of ketones to carbinols by means of fermenting yeast is completely different from the method of resolution of racemic alcohols by treatment with living microorganisms (Pasteur). In the latter case one of the enantiomorphs is removed by oxidation during metabolism in the former it is produced by true asymmetric hydrogenation, without the intermediate formation of the inactive form, (Cf. Mayer and Levene and Walti. )... 

The oxidation of a secondary alcoholic group in the presence of primary alcoholic groups by Acetobacter suboxydans converts adonit into adonose [1041]. The treatment of androstenediol and dehydroandrosterone with yeast yields A -androstenedione [1088]. Steroidal hydroxy ketones are dehydrogenated to dicarbonyl compound with CorynebacteriUlh simplex [1056] (see equations 446 and 447). Examples of oxidations of secondary alcohols to ketones are shown in equations 265-268. 

Cha et al. provided the first experimental proof of hydrogen tunneling on an enzyme by reporting an elevated RS exponent for benzyl alcohol oxidation by yeast ADH (YADH) [10]. Isotope effects for benzyl alcohol oxidation were determined by the mixed-label tracer method, in which the primary and a-secondary positions of benzyl alcohol are either H or D, with stereochemically random, trace-level T incorporation. In this fashion, the observed ratios between the a-secondary (kH/feT)i H and (kD/feT)i°D KIEs are susceptible to both Swain-Schaad and RGM deviations and, thus, are sensitive probes for tuimeling (see Section 10.3.3.3). The observed a-secondary RS exponent, kn/feT = at 25 "C, greatly exceeded... 

In the case of 1,3-butanediol, the remaining diol had slight optical activity. This showed (S)-configuration at 27% opticd purity. This was determined by comparison of optical rotation with a reference (6). The yeasts could differentiate the (R)- and (S)-alcohol and oxidize the (R)-dcohol predominantly. With 1,4-pentanediol, the yeasts preferred the primary hydroxy group to the secondary one and gave pentane-1,4-olide and its lactol at 6.3% yield. The afforded lactone showed slight optical activity and was the (R)-form at 5% optical purity. This was determined by comparison of optical rotation with a reference (7). This showed that the yeasts preferred (R)-l,4-pentanediol as the substrate. 

Yeast alcohol dehydrogenase (YADH), catalysis of reduction by NADH of acetone formate dehydrogenase (FDH), oxidation by NAD of formate horse-liver alcohol dehydrogenase (HLAD), catalysis of reduction by NADH of cyclohexanone With label in NADH, the secondary KIE is 1.38 for reduction of acetone (YADH) with label in NAD, the secondary KIE is 1.22 for oxidation of formate (FDH) with label in NADH, the secondary KIE is 1.50 for reduction of cyclohexanone (HLAD). The exalted secondary isotope effects were suggested to originate in reaction-coordinate motion of the secondary center. 

In the following year, Cleland and his coworkers reported further and more emphatic examples of the phenomenon of exaltation of the a-secondary isotope effects in enzymic hydride-transfer reactions. The cases shown in Table 1 for their studies of yeast alcohol dehydrogenase and horse-liver alcohol dehydrogenase would have been expected on traditional grounds to show kinetic isotope effects between 1.00 and 1.13 but in fact values of 1.38 and 1.50 were found. Even more impressively, the oxidation of formate by NAD was expected to exhibit an isotope effect between 1.00 and 1/1.13 = 0.89 - an inverse isotope effect because NAD" was being converted to NADH. The observed value was 1.22, normal rather than inverse. Again the model of coupled motion, with a citation to Kurz and Frieden, was invoked to interpret the findings. 

The electron transfer between NADH and the anode may be accelerated by the use of a mediator. Synthetic applications have been described for the oxidation of primary and secondary alcohols to aldehydes and ketones catalyzed by yeast alcohol dehydrogenase (YADH) and the alcohol dehydrogenase from Thermoanaerobium brockii (TBADH) with indirect electrochemical regeneration of NAD+ and NADP+, respectively, using the tris(3,4,7,8-tetramethyl-l,10-phenanthroline) iron(II/III) complex as redox catalyst [59],... 

ADH features another catalytic triad, Ser-Tyr-Lys. Whereas the liver ADH kinetic mechanism is highly ordered, coenzyme associating first and dissociating last, the yeast ADH mechanism is largely random. In both cases, the actual chemical reaction is a hydride transfer. In the oxidation of secondary alcohols by Drosophila ADH (DADH), the release of NADH from the enzyme-NADH complex is the rate-limiting step, so vmax is independent of the chemical nature of the alcohol. With primary alcohols, as vmax is much lower and depends on the nature of alcohol, Theorell-Chance kinetics are not observed and the rate-limiting step is the chemical interconversion from alcohol to aldehyde. 

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