© 2006 The Authors Journal compilation © 2006 The Royal Microscopical Society Journal of Microscopy, Vol. 224, Pt 3 December 2006, pp. 290–297 Received 09 December 2005; accepted 11 July 2006 Blackwell Publishing Ltd Digital image processing as a tool to monitor biomass growth in Aspergillus niger 3T5B8 solid-state fermentation: preliminary results S. COURI, E. P. MERCÊS, B. C. V. NEVES* & L. F. SENNA† Embrapa/Agroindústria de Alimentos, Av. das Américas, 29.501 Guaratiba, Rio de Janeiro/RJ, Brazil 23020-470 *Universidade Federal Rural do Rio de Janeiro, Departamento de Tecnologia de Alimentos, Seropédica/ RJ, Brazil †Universidade do Estado do Rio de Janeiro, R. São Francisco Xavier, 524 Pav. Haroldo Lisboa, S. 427 Maracanã/RJ, Brazil Key words. Biomass growth, digital image processing, filamentous fungi, glycosamine analysis, polygalacturonase, solid-state fermentation. Summary The estimation of biomass is an essential parameter for controlling fermentation processes. However, monitoring biomass growth in filamentous fungi solid-state fermentation is laborious. The aim of this study was to provide a better insight into the monitoring of biomass growth in Aspergillus niger 3T5B8 solid-state fermentation using a digital image- processing technique. The images were acquired with a stereomicroscope and a digital camera, and processed using KS400 software. Growth was evaluated every 24 h for 5 days, and quantified as the total area occupied by the hyphae. The correlation between the results of the proposed methodology and the polygalacturonase data was greater than 0.9, showing that a direct and linear relationship can be expected among these parameters. This work indicates that the digital processing technique can be used for indirect biomass estimation in a solid-state fermentation process. Introduction The design and operation of a fermentation process can be improved by using methodologies for biomass estimation that permit investigation of the relationships among growth kinetics, biomass morphology and the fermentation product (Tucker et al., 1992; Loera & Viniegra-González, 1998). With solid-state fermentation (SSF), it is harder to obtain accurate estimates, mainly because of the difficulties of biomass– fermentation medium separation, but also owing to the dependence of the obtained data on the biomass properties and on the methodology parameters, such as sensitivity and estimation speed. In fact, very few data are available concerning the kinetic and morphological characterization of biomass in SSF, mostly because of the complexity of measurement of the main process variables. Some methodologies have been proposed as indirect measurements of biomass quantification, such as the production of primary metabolites (Okazaki et al., 1980; Raimbault, 1998; Neves, 2003) or carbon dioxide (Carrizalez et al., 1981), the variation in the electrical conductivity between biomass and the solid substrate (Peñaloza et al., 1991), the changes in the colour of the fermentation medium, determined by using reflected light (Murthy et al., 1993), the quantification of intracellular protein (Favela-Torres et al., 1998) or other compounds such as ergosterol (Desgranges et al., 1991), and enzyme-like immunosorbent assays, which quantify antibody reactivity in the mycelium cell wall (Dubey et al., 1998). In most cases, the proposed methodology was compared to the glycosamine method, revised by Penman et al. (2000). This is a classic method for biomass quantification, where the quitine monomer present on the fungi cellular wall, N-acetyl-d- glycosamine, is measured. However, this method can present errors due to the variation in the glycosamine content found at different growth intervals, and the low specificity of its quantification. This means that different six-carbon sugars present in vegetal tissues, such as galactosamine, can also be Correspondence to: S. Couri. Tel: +55 21 24109618; fax: +55 21 24109616; e-mail: scoury@ctaa.embrapa.br Received 09 December 2005; accepted 11 July 2006