Optimizing red sorghum malt quality whenBacillus subtilisis used during steeping to control mould growth

Abstract

Previous work has shown that Bacillus subtilis-S499-based biocontrol treatments applied without aeration at the steeping stage of red sorghum malting offer good mould reduction, but yield malts with low levels of key hydrolytic enzymes. Thus we attempted to raise these levels by aerating the steeping liquor, varying the steeping time (from 8 to 40 h) and temperature (from 25 to 35 C), and combining a biocontrol treatment with prior steeping in 0.2% NaOH. Aeration proved particularly important whenever B. subtilis cells were present in the steep liquor. The optimal temperatures for a-and b-amylase were 30 and 25 C, respectively. By increasing the steeping time, it was possible to improve the a-amylase activity, but the b-amylase activity peaked sharply between 16 and 20 h, depending on the steeping medium. A good compromise was steeping in a biocontrol medium for 14-16 h at 30 C. Combination steeping treatments (0.2% NaOH for 8 h followed by biocontrol for 8 h) yielded malts of a quality approaching that afforded by dilute alkaline treatment. Copyright © 2012 The Institute of Brewing & Distilling Supporting information can be found in the online version of this article.Keywords: red sorghum; biocontrol; mould reduction; malt quality; amylases; Bacillus subtilis IntroductionIn brewing, mould contamination of the grains used to produce malt can lead to the introduction of mycotoxins into the brewing process. Mycotoxins constitute a health hazard for consumers (1-3). They can also affect the brewing yield (4), the physiological state of the yeast (4) and beer quality (5). Moulds and their mycotoxins are thus an expensive problem for the malting and brewing industries (6). This concern exists in the case of barley malting [see Dodd (6), for instance], but it is particularly acute in the case of sorghum malting. One reason is that sorghum grains have no protective husks and are therefore more vulnerable than barley to mould contamination owing to grain damage. A second reason is that, in many parts of Africa and Asia, traditional outdoor floor malting under uncontrolled conditions is the rule. Thirdly, whether traditional or industrial, the steeping and germination steps of sorghum malting require both high humidity and a relatively high temperature [ideally, about 30 C (7) vs 12-18 C for barley], and are thus highly favourable to microbial development.As an alternative to the use of toxic chemicals such as formaldehyde, steeping in a dilute alkaline solution (0.1-0.5% NaOH or CaOH 2 ) has been found to reduce the level of mould contamination without adversely affecting the sorghum grain (8,9). In addition, it can improve the quality of the malt, notably by inactivating polyphenols/ condensed tannins and increasing both the diastatic power (DP) and the free amino nitrogen (FAN) (7,(10)(11)(12). Biocontrol strategies are also envisaged, that is, the use of harmless microbes to control the fungi that produce mycotoxins and gushing-promoting agents (13-15). Lefyedi and Taylor (13) have tested the effect of steeping in th...

Modelling the β-amylase activity during red sorghum malting when Bacillus subtilis is used to control mould growth

“…Congo for producing alcoholic beverages and has been described previously (Bwanganga et al, 2011(Bwanganga et al, , 2012. It contains: 3.8% AE 0.3 fat content, 2.9% AE 0.1 b-glucans, 73.9% AE 1.9 total starch (as assayed by the Megazyme K-TSTA 07/11 method), 27.6% AE 1.4 amylose (assayed by the Megazyme K-AMYL 07/11 method), 8.1 mg Gallic Acid Equivalents per gram of dried malt (assayed using the method optimized by Georgé et al (2005) without eliminating water-soluble compounds) and 0.18% Cathechin Equivalent (assayed using the modified vanillin method of Price et al (1978)).…”

“…It contains: 3.8% AE 0.3 fat content, 2.9% AE 0.1 b-glucans, 73.9% AE 1.9 total starch (as assayed by the Megazyme K-TSTA 07/11 method), 27.6% AE 1.4 amylose (assayed by the Megazyme K-AMYL 07/11 method), 8.1 mg Gallic Acid Equivalents per gram of dried malt (assayed using the method optimized by Georgé et al (2005) without eliminating water-soluble compounds) and 0.18% Cathechin Equivalent (assayed using the modified vanillin method of Price et al (1978)). Sorghum malts were obtained as described by Bwanganga et al (2012) by manual sorting, aerated steeping at 30 C for 8 h in 0.2% NaOH followed by 8 h resteeping in the biocontrol steep liquor (distilled water containing 10 8 , 10 4 B. subtilis S499 cells (respectively for B1 and B2), distilled water alone being used as a control (C)); germination in the dark (to exclude photosynthesis) at the indicated temperature (25, 30, or 35 C) for the indicated time (6, 12, 24, 36, 48, 60, 72, 84, 96, 108, 120, 132, 144, 156, 168 and 180 h), and kilning for 48 h at 40 C. Beta-amylase activities were extracted and assayed using Megazyme methods Betamyl-3 (K-BETA3 10/10). Sorghum b-amylase was extracted using 0.05 M Tris-HCl buffer plus 1 mM EDTA, dilution was made in 0.1 M MES buffer plus 1 mM EDTA, 1.0 mg/ml of BSA and 0.02% w/v sodium azide.…”

“…Yet the malts obtained showed lower amylase levels than the malt steeped at the same temperature and for the same length of time in 0.2% NaOH, a treatment known to yield sorghum malts of good quality (Beta et al, 2000;. In a second study (Bwanganga et al, 2012) we sought to optimize the biocontrol treatment in terms of malt quality. The best results were obtained by aerating the steep liquor and replacing the 16 h biocontrol treatment with 8 h steeping in 0.2% NaOH followed by 8 h resteeping in the B. subtilis S499-based biocontrol treatment.…”

“…In an initial study (Bwanganga et al, 2011), steeping for 16 h at 30 C in the presence of a diluted 72 h B. subtilis S499 culture (cells, medium, or cells þ medium) was found to reduce the level of fungal contamination in the kilned malt. Yet the malts obtained showed lower amylase levels than the malt steeped at the same temperature and for the same length of time in 0.2% NaOH, a treatment known to yield sorghum malts of good quality (Beta et al, 2000;.…”