PRODUCTION Of NUTRIENT SOURCES FOR RHIZOBIUM

mannitol, xylitol, and sorbitol were used as standards for the comparison of R f.. The standards gave R f
0.741, 0.464, 0.433, 0.406, and 0.350 for glycerol, xylitol, glucose, mannitol, and sorbitol
respectively, which corresponded to that of authentic sugar alcohols.
In culture broth, 117 isolates produced glycerol could be detected on TLC chromatogram, and
3 isolates could produced high concentration of mannitol on chromatograms. In cell lysate, the
glycerol spot were found slightly from extracts of 2 isolates, and 13 isolates produced mannitol (Table
10). For 15 type strains, the culture broth of Saccharomyces cerevisiae var. burgundi, S. cerevisiae
var. champaign, H. anomala TISTR 5082, K. maxianus TISTR 5270, and S. baryanus showed dense
spots of glycerol on chromatograms. Saccharomyces cerevisiae, C. utilis, C. famata, C. krusei, and C.
tropicalis, and S. fibuligera TISTR 5067 could also produce glycerol detected from their culture broth
(Table 11, Figure 14, 15 and 16). In cell lysate, only Rhodotorula rubra TISTR 5067 produced
mannitol detected on chromatogram (Figure 12 and 13). TLC chromatograms of yeast culture broth
and cell lysates were shown in Figures 12-16. Mannitol was detected from 11 isolates (AFY4, LLY3,
AFY5, SLY1, TAY7, WAY6, KAY1, TAY9, PIY2, PUY4, and Y60) and 1 type strain, R. rubra
TISTR 5067 (Figure 12 and 13). Glycerol was detected 9 isolates (COY1, SWY1, BLY2, APY6,
FAY1, FAY2, LIY2, TAY9 and CIY1) (Figures 14, 15, and 16).
The Glycerol combination test kit was also used for the determination of glycerol in the
screening step. Thirty-three yeast isolates and 1 type strain, K. maxianus TISTR 5270, showed dark
spots on TLC chromatograms were selected for quantitative analysis of glycerol. Nine yeast isolates
with the four scores (Table 10) gave glycerol productivity approximate 1.86-4.64 g/l. The yeast isolate
LIY2 gave the highest glycerol production, 4.64 g/l. The mannitol production was quantified by using
High-Performance Liquid Chromatography (HPLC). Three yeast isolates (KAY1, PIY2, and PUY4)
produced both of mannitol and glycerol in yeast cells and could also produced in cultured medium. At
the three scores, the highest glycerol production were 2.66 g/l and 1.68 g/l respectively from yeast
isolate APY6 and K. maxianus TISTR 5270 (Figure 17).
For the determination of glycerol and mannitol using High-Performance Liquid
Chromatography (HPLC), the sugar alcohols production was compared with the spot on
chromatogram. Yeast isolates KAY1, PIY2, and PUY4, and R. rubra TISTR 5067 could produce
glycerol and mannitol in both cultured media and cell lysates (Table 12). In cultured medium, the
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