The thermotolerant, ethanol-producing yeast strain, Kluyveromyces marxianus IMB3, has been immobilized in calcium alginate matrices. The ability of the biocatalyst to produce ethanol from cane molasses originating in Guatemala, Honduras, Senegal, Guyana and the Philippines was examined. In each case the molasses was diluted to yield a sugar concentration of 140 g/l and fermentations were carried out in batch-fed mode at 45°C. During the first 24 hours, the maximum ethanol concentrations obtained ranged from 43-57 g/l with optimum production on the molasses from Honduras. Ethanol production during subsequent re-feeding of the fermentations at 24-hour intervals over a 120-hour period, decreased steadily to concentrations ranging from 20-36 g/l and it was found that ethanol productivity remained highest in fermentations containing the molasses from Guyana. When each set of fermentations was re-fed at 120 h and allowed to continue for 48 h, ethanol production again increased to a maximum with concentrations ranging from 25-52 g/l. It was also found however, that increasing the time between re-feeding at this stage in fermentation had a detrimental effect on the functionality of the biocatalyst.

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The thermotolerant ethanol-producing yeast strain Kluyveromyces marxianus IMB3 was immobilized in calcium alginate and a 1:1 mixture of calcium alginate and the porous volcanic mineral, kissiris. Immobilized preparations were placed in ®xed-bed column bioreactors and continuous ethanol production by systems containing both immobilized preparations was examined at 45°C with a 100 g/l glucose feed. The effect of residence time on product concentration, bioreactor ef®ciency and volumetric productivities have been examined and these were all higher in systems containing the alginate/kissiris mixed immobilization matrix. Maximum ethanol concentrations produced by the continuous system ranged between 46 and 48 g/l representing ef®ciencies of 90±94%. IntroductionThe thermotolerant yeast strain, Kluyveromyces marxianus IMB3, has been shown to produce ethanol during growth on carbohydrate sources such as glucose [1], sucrose [2] and lactose [3] at 45 o C. As a result of its ability to perform this function at relatively high temperatures this yeast is ideally suited for use in simultaneous sacchari®-cation and fermentation (SSF) processes and it has been shown to be capable of ethanol production on substrates such as starch and cellulose [4,5]. In addition to the above advantage associated with the ability of this yeast to function at 45°C or indeed higher, other advantages include a reduced risk of bioreactor contamination and increased ef®ciency with respect to product formation and recovery. In a series of recent studies our group has shown that it is possible to immobilize this yeast in alginate-based matrices and produce ethanol in batch-fed con®gurations from sugars such as glucose, sucrose and lactose [6,7,8]. It has also been demonstrated that these immobilized preparations may be incorporated into simultaneous sacchari®cation and fermentation processes concerned with the conversion of cellulosic materials to ethanol [9]. In many of the above studies the immobilized biocatalyst preparations were much more stable than the free microorganism, particularly over prolonged periods of time. In addition, higher ®nal concentrations of ethanol were produced by the immobilized preparations over longer periods of time in continuous batch-fed systems [8]. Although a variety of con®gurations involving semi-continuous batch-fed systems have been described for ethanol production by K. marxianus IMB3, the major objective of our group has been to apply continuous-¯ow technology to ethanol production by this thermotolerant microorganism.Although alginate is a commonly used immobilization matrix in laboratory studies [10] and provides an ideal and acceptable means of studying a variety of model systems at that level, the economic viability of any ethanol production process would depend heavily on biocatalyst preparation costs. Reducing the quantity of alginate in alginate-based immobilization biocatalyst formulations would be expected to contribute positively to the economics of bioconversion. In recently reported studies it has bee...

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