Trends in Food Science & Technology 14 (2003) 109110

Letter

Problems with enzymic bioprocessing of foodstus

Alan Wiseman
c/o Biochemistry Group, School of Biomedical and Life Sciences, University of Surrey, Guildford GU2 7XH, UK

We have documented the positive side of the use of soluble enzymes and immobilized enzymes (IME) (Tucker & Woods, 1997; Wiseman, 1995) over a 40 year period: although we are aware that enzymes were used in the 19th C to make malt loaf by the use of diastase (-amylase), largely as malted barley extract to convert wheat starch into maltose, in situ in the bread-dough prior to baking. Moreover, immobilized sucrases (invertases) from yeast, or the yeast itself, was used before the turn of the 20th Century to make invertsugar (golden syrups). Enzymic bioprocessing of foodstus (Tucker & Woods, 1997; Wiseman, 1995) on large-scale can be exemplied with amylases use in bread-making. In a dry year, the USA practice was to supplement amylasedecient wheat our; by addition of sucient fungal amylase to provide glucose to bakers yeast (Saccharomyces cerevisiae), added later. Raising of the dough was therefore achieved and at least 1015 advantages of supplementation were noted in the production of, and composition of, the marketed bread: these claims have included improvement in the bread digestibility, texture, crust colour, staling resistance, absence of crumb stickiness along with enhanced sweetness. Nevertheless, enzymic over-treatment lowers the saleability of the various dough-bakes. Especially worthy of note is the limited hydrolytic ability of Ca2+-dependent fungal amylases, that especially is quickly terminated in

the hot oven unit-operation (unlike the corresponding relatively thermostable microbial a-amylases). In brewing practices to dissolve wheat-derived adjuncts to supplement the malted barley (up to 80% adjunct to 20% barley-malt), from largely traditional malting, there can be use also of added gibberellin (plant-hormone) to accelerate the barley to malt bioconversion. Moreover, a two-step enzymic bioprocessing (amylolytic/proteolytic) takes only a few hours at appropriate moderate temperatures, and any perceived changes in beer (ale) avour and fragrance have not been viewed as consumer-unfriendly (unlike some 1960s earlier continuous-fermentation brews). Beer itself can be viewed as an enhanced-multifunctional food (Wiseman & Woods, 2001); because of its partially yeastderived vitamin and mineral content. Furthermore, the ethanol content may increase the bioavailability (due to g.i. tract bioconversions by microora there) of nutraceuticals such as particular polyphenols (Pfannhauser, Fenwick, & Khokar, 2001) so that multifunctional health benets ensue. In addition, any dysfunctionality due to higher alcohols (fusel oils) may become minimized; either by enzyme treatment or by inhibition of partitioning-uptake ability of gut epithelial cells. Furthermore, penetration of the bloodbrain barrier (mounted by neuroglial cells, in prevention of neuronaluptake of harmful food components) can ensue. For instance l-glutamate (the precursor of -aminobutyric acid inhibitory-transmitter: GABA) causes the symptoms from monosodium glutamate consumption in some individuals. Moreover, l-tryptophan, manufactured to supplement foods decient in this essential amino acid, can be too readily converted to the brain transmitter serotonin (5-hydroxytryptamine, 5-HT) and may need to be removed using enzymes, where individual nutrigenomic contraindications are identied. Enzymic treatment suers from a variety of problems caused by poor thermostability of some natural enzymes unless immobilised (Wiseman & Woods, 2002b). Nevertheless, the ease of rapid curtailment of biocatalysts by heating is often essential: the critical temperature (e.g. 65 C) is where there is the collapse of enzymic active conformation in thermal denaturation (often with unfolding of the enzyme). Such denaturation is usually irreversible although there is evidence that restoration of enzyme activity can be achieved by the use of hydrophobic-interaction lowering agents such as

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Alan Wiseman / Trends in Food Science & Technology 14 (2003) 109110

urea (this restoration of activity needs slow incubation) or hydrogen bond-breaking agents such as guanidinium hydrochloride. Understanding now is emerging of prion-protein unfolding energetics (Dobson, 2002) followed by britic-aggregation perhaps with conformation change as that of Cu-prion, with superoxide dismutase ability in activation of the brain-disturbing form (Wiseman & Woods, 2002b). Similarly, denaturation kinetics of food proteins also will become predictable in silico, to the benet of ecacious food bioprocessing with enzymes and immobilised enzymes (Wiseman & Woods, 2002a). Such enzymic processes have suered from notable take-up reluctance (over a 30 year period); this may be because of fears of heavy metals causing unpredictable inhibitor-initiated failure in the planned bioprocessing. Nevertheless, these metals can be readily removed by high-anity binding proteins, such as the metallothionins. Brewers yeast, now genetically dened, although a multifunctional food with high vitamin and mineral content, is best known at present for genomic-proling studies (Glaver et al., 2002) with explanations inter alia of high NaCl (1M) growth-phenomena relative to gene expression changes (these were poorly correlated). Brewing produces edible yeast of high quality free from toxic heavy metals (and is suitable for NaCl or thermal autolysate production), and carbon dioxide (for carbonation of soft drinks, alcoholic or otherwise, and for liquid CO2 solvent extraction in, for example, decaeinated beverages production). Moreover, brewers yeast holds invertase (and acid phosphatase) in the periplasmic space; inside the cell wall, where however, it is fully accessible to sucrose: and can be used therefore in immobilized cell (IMC) processes as a substitute for IME. Moreover, invertase is a glycoprotein and although highly thermostable is subject to lectin (e.g. concanavalin

A) liganding inhibition: although lectin can be destroyed by deployment of specic enzymes. Cautionary tales of problems with bioprocesses that utilise such glycoprotein enzymes abound (partially due to poor denition of the eect of the glyco-moiety): these include aberrant running in various types of electrophoresis (and in gel-exclusion dened pore-size chromatography). In silico redesigned enzymes and mimics are only now emerging, but their toxicity remains to be assessed (Wiseman et al., 2000). This is especially important with selenoproteins, because we require 50 mg per day in our diet: whereas consumption of 500 mg per day is toxic (Rayman, 2002).

References
Dobson, C. (2002). Getting out of shape Nature, 419, 729730. Glaver, C., Chu, A.M., Ni, L., Connelly, C., and Rules, L. et al. (2002). Functional proling of Saccharomyces cerevisiae genome. Nature, 418, 387391 Pfannhauser, W., Fenwick, G. R., & Khokar, S. (2001). Biologicallyactive phytochemicals in food: analysis, metabolism, bioavailability and function. London, UK: RSC. Rayman, M. (2000). Se brought to earth Chemistry in Britain, 38, 28 31. Tucker, G., & Woods, L. F. J. (1997). Enzymes in food processing (2nd ed.). Glasgow, Scotland, UK: Blackie & Sons. Wiseman, A. (Ed.). (1995). Handbook of enzyme biotechnology (3rd Ed.). Hemel Hempstead, UK: Ellis Horwood (Prentice Hall). Wiseman, A., & Woods, L. F. J. (2001). An enhanced multifunctional foodbeer? The BREWER International, 1, 55. Wiseman, A., & Woods, L. F. J. (2002a). Precautions against immobilised enzyme (IME) inhibitors Trends in Food Science & Technology, 12, 469470. Wiseman, A., & Woods, L. F. J. (2002b). Welcome praise for copper Chemistry in Britain, 38, 20. Wiseman, A., Lewis, D. F. V., Ridgway, T. J., & Wiseman, H. (2000). Cytochromes P-450 in food-processing: limitations of imitations Journal of Chemical Technology and Biotechnology, 75, 35.

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