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Various microorganisms including bacteria, yeast and fungi can degrade caffeine. There are few publications about caffeine degradation pathway in filamentous fungi, mainly by solid-state fermentation (SSF). Studies were carried out on degradation of caffeine and their metabolites by filamentous fungi in SSF using coffee husk as substrate. The purpose of this work was to investigate the caffeine degradation pathway by Rhizopus delemar in packed bed column fermenter and to compare this degradation metabolism with glass flasks fermentation. The methylxanthines were quantified by HPLC analysis. The experiments were realized with the optimized conditions in previous experiments: pH 6.5, 28ºC, inoculation rate 10 6 spores/g substrate, aeration rate 60 mL/min and initial moisture 73%. Under these conditions, after 72 hous of fermentation was achieved only 0.19% of caffeine and 0.014% of theophylline in the coffee husk. The strain proved to be able for caffeine and theophylline degradation by SSF in packed bed column bioreactor.

Advances in biotechnology, mainly in the area of enzymes and fermentation technology, offer potential opportunities for economic utilization of agro-industrial residues. The coffee husk obtained during dry processing of coffee has been poorly utilized when compared with the produced volume. Removing the toxics compound as caffeine and tarmin, this residue could be more utilized. Filamentous fungi are able to assimilate caffeine and tannins :from liquid synthetic media or coffee residue. There are few works about coffee husk detoxi.fication and it was never described with strains of Rhizopus delemar. The caffeine degradation pathway and the enzymes involved in this process have not been described in the literature to. The aim of this work was to improve the nutritive value of coffee husk by solid-state fermentation with R. delemar, to investigate the caffeine degradation pathway and to characterize enzymes involved in this bioprocess. The conditions of coffee husk fermentation by R. delemar were optimized and the best detoxi.fication leveis (86% of caffeine and 60% of tannin reduction) were obtained in Erlenmeyer flasks with 75% initial moisture, I 0 6 spores/g dry substrate, at pH 6.5 and 28°C.The kinetic o f fermentative process was carried out in packed bed column bioreactor using the conditions previously optimized and showed that the development o f filamentous fungi and its respiration was related with caffeine degradation and total sugars presents in coffee husk. The exponential phase o f microbial growth was achieved between 3 and 5 days, when was detected maximal Oz consumption and after the maximum production of enzymes and caffeine degradation. The maximum activities, 1.8 and 3.3 U/g d.s. for caffeine and theophylline demethylase respectively, were achieved at 2 days of fermentation in Erlenmeyer flasks. The enzymatic extracts were stable after storage at 2°C and the Km estimated, the optimum temperature and pH were respectively 150J.LM, 30°C and pH 6.5 for caffeine demethylase and 180J.LM, 30°C and pH 7.4 for theophylline demethylase. The results showed good prospects ofusing Rhizopus sp. for the coffee husk detoxi.fication and would open new opportunities for the utilization of this byproduct and for enzymatic extract application.

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