Introduction and key issues

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The history of food processing technologies is as long as the history of mankind itself. Our ancestors used different techniques to preserve and convert foods. One of the most important conversions is biocatalysis using enzymes or cells in food processing. From a historical perspective, acid proteases have been considered as the oldest enzymes used (Hofmann, 1974). One isolated enzyme of the group - chymosin, in the form of rennet - has been used for thousands of years. People in the twenty-first century - being more and more concerned about environmental problems

- intend to use these 'ancient' methods not only in the processing of foods, but also in waste management and co-product recovery in the food industry. Great efforts have been made to find and develop proper and effective biocatalytic techniques in this area. Since biocatalysis - in general

- is a mild, highly selective and environmentally safe tool in processing technologies, its application is sensible in handling wastes and co-products formed in the food industry. In addition to these key reasons, it is important to note that in many cases the methods involving enzymes and microbial cells are waste-free and produce (sellable) value-added compounds, while solving the environmental problems caused by wastes and co-products. Thus biocatalytic methods can contribute significantly to higher effectiveness in food technologies (Birch et al., 1981; Tucker and Woods, 1998).

Enzymatic extraction and fermentation 199 9.2 Biocatalytic methods

The application of biocatalytic methods to the handling of waste and co-products coming from food technologies can be divided into two main groups:

• methods without product formation;

• methods with product formation.

Firstly there are procedures existing where no product is formed during the treatment of solid waste or waste water from food processing. In these cases, degradation of acid content or simply reduction of the waste's volume are the main purposes (Elmaleh et al., 1999). Similarly, preservation - and not product formation - may be the aim of ensilation processes, e.g. shrimp waste ensilation by lactic acid bacteria (Shirai et al., 2001). On the other hand, product(s) may be obtained from the waste or co-product; this is considered to be a more sophisticated, enhanced method. Energy and/or any kind of material (compounds) can be produced in this way. The energy production can be realised by, for example, microbial fuel cells (Grzebyk and Pozniak, 2005; Ieropoulos et al., 2005). This kind of equipment, however, currently only exists at laboratory scale and the scientific fundamentals are being studied in detail; thus, its application is expected in the next decade. Biogas production - as an energy source - on the other hand, is widely realised by anaerobic fermentation, using mixtures of microbe populations. However, this method is not restricted to waste management in the food industry and is therefore not discussed here.

The production of various compounds from food wastes and co-products by biocatalytic methods has been investigated for a long time and many interesting and surprising examples can be listed. Some of these are presented below to illustrate the extremely wide spectrum of uses; these uses are arranged according to the particular product of the fermentation and enzymatic procedures. Although a number of different products, biocata-lysts and methods are given here, the list is far from complete.

9.2.1 Fermentation

Single cell protein (SCP)

SCP is the dried form of various microorganisms, such as bacteria (sometimes called bacterial protein), fungi and algae grown on different kinds of substrates. SCP production on solid as well as liquid food wastes is one of the well-known fermentation processes. Numerous raw materials are suitable for the process (Ozyurt and Deveci, 2004), e.g. molasses (Nagy, 1998), starch processing waste water (Jin et al., 1998), lemon pulp and peel (De Gregorio et al., 2002) or whey (Moeini et al., 2004) are equally appropriate initial substances. In the procedure based on beet molasses, Saccharomyces cerevisiae yeast is applied and pure oxygen is supplied which prevent bacterial multiplication as well as foam formation. The protein content of the SCP is approximately 70%. Aspergillus oryzae was selected for SCP fermentation from starch waste, while Aspergillus niger and Trichoderma viride species were applied in the slurry-state fermentation of citrus pulps. Using whey as a substrate, a mixed yeast culture was found suitable and effective for improved SCP production.

In certain cases it is also possible to simultaneously produce enzymes during SCP production. An interesting example of this was a study where extracted grape waste material and pressed apple pulp were tested as carbon sources for growing Penicillium funiculosum, Myrothecium verrucaria and Aspergillus niger, with the aim of producing cellulolytic enzymes (Kuzmanova et al., 1991). Crude protein (35%) and a high level of cellulase activity was produced during the test.

Production of enzymes

A wide variety of enzymes are produced by the fermentation of solid and/or liquid wastes and co-products. Table 9.1 shows the range of hydrolytic enzymes, the microorganism applied and the food waste/co-product used. In the class of hydrolases (EC 3), amylases, cellulases and hemicellulases, pectinases, proteases and lipases are the most important enzymes in the processing of various food wastes. The feed mixture of these processes would usually contain the main compound(s) targeted for degradation or convertion by the enzyme. For example, in the case of pectinase enzymes, a pectin-containing mixture is required as feed in the fermentation (enzyme induction).

The majority of these hydrolytic enzymes act on O-glycosyl compounds (EC 3.2.1) and are used for degradation of polysaccharides (cellulose, starch, xylanes, pectin, chitin), while lipases (EC and peptidases (EC 3.4) hydrolyse carboxylic ester bonds (triglycerides) and peptide bonds (proteins), respectively. These substances occur in almost all types of wastes and co-products from food processing. Therefore their degradation is extremely important and highlights the need for commercial quantities of these enzymes. The most cost-effective way to obtain these enzymes is to synthesise them 'on-site', using the same substrate; i.e. simultaneous conversion of biomass and the production of biocatalysts is undoubtedly beneficial, moreover these systems allow much higher flexibility in processing (Gao et al., 2002). The other benefit is the possible adaptation of the species to the particular substrate, resulting in higher enzyme activity (Juhász et al., 2005).

Food wastes and co-products are also suitable materials for the production of other (non-hydrolytic) important industrial enzymes. A few examples are: fumarase (EC and aspartase (EC production on molasses by Erwinia species for the biotransformation of fumaric acid into L-malic acid and L-aspartic acid, respectively (Bagdasaryan et al., 2005); laccase (EC produced by Botrytis cinerea (Howard et al., 2003);

Table 9.1 Production of hydrolytic enzymes on waste or co-products from the food industry



Waste/ co-product

Reference a-Amylase a-Amylase a-Amylase Amylases

Cellulases Cellulases Hemicellulases Xylanases



Pectinases Lipase


Acid proteases

P-N-Acetylhexos-aminidase (chitinase)

Bacillus coagulans

Bacillus subtilis

Botryodiplodia theobromae Rhizopus oryzae Phanerochate chrysosporium Trichoderma reesei e.g. Botyris cinerea Trichoderma harzianum

Aspergillus niger

Aspergillus carbanerius Thermoascus aurantiacus

Aspergillus niger

Penicillium simplicissimum

Penicillium restrictum

Verticillium silage lecanii

Wheat bran

Banana fruit stalk

Cellulomonas sp. Wheat bran

Cassava starch residue Soy hull

Rice straw

Wheat straw

Rice/wheat straw, sugarcane bagasse Sugar beet

PulP Wheat bran

Orange bagasse Sugar cane bagasse Wheat bran Apple pomace

Babassu cake, sugarcane molasses, corn steep liquor Babassu oil cake Surimi wash water Shrimp waste

Babu and


1995 Krisha and



Emtiazi and Nahvi,

2004 Ray, 2004

Kaur et al., 1998

Thygesen et al.,

2003 Abdel-Sater and El-Said, 2000

Naidu and Parda, 1998

Martins et al., 2002

Berovic and Ostroversnik, 1997 Gutarra et al., 2005

Gombert et al.,

1999 DeWitt and

Morrissey, 2002 Matsumoto et al., 2004

lignin peroxidase (EC and manganese peroxidase (EC production by Phanerochaete chrysosporium on lignocellulose wastes (e.g. straw) for removal of toxic phenolic compounds from industrial waste water (Fujian et al., 2001).

Acid compounds

Various acid compounds - used as acidifying agents in the food industry -are synthesised based on food wastes or co-products. Two of the most important are citric acid and lactic acid. Citric acid was manufactured, for example, by Aspergillus niger on apple pomace in a packed bed bioreactor (Shojaosodati and Babaeipour, 2002), but it was found that ram horn pepton is also a suitable protein source using the same strain (Kurbanoglu and Kurbanoglu, 2004). Lactic acid production from the dairy co-product whey is a 'traditional' technique, where lactose content is converted by bacteria species (Nagy, 1998). Glucose is similarly a suitable substrate for lactic acid formation (Tucker and Woods, 1998). Lactic acid may also be produced on canned pineapple syrup, by Lactococcus lactis, applying grape invertase enzyme to improve the utilisation of sucrose from the syrup (Ueno et al., 2003). Starch-containing food wastes are also applicable for lactic acid production, although saccharification of starch is required beforehand. Saccharification and fermentation may be achieved simultaneously using amylolytic enzyme preparations and, for example, Lactobacillus delbrueckii (Kim et al., 2003).

Biodegradable polymers

Biodegradable polymers (Smith, 2005) are possible products in glycerol processing (glycerol is the by-product of, for example, fat splitting or biodiesel production). Among these green polymers, an important group are the polyesters, including the polyhydroxy alkanoates (PHAs) and polyesters of 1,3-propanediol (PD) which can be produced not only from glycerol, but also from other food wastes and co-products. In the case of PHAs, the length of the alkyl chain may be varied but butyrate is the most favourable compound. Its fermentation (Table 9.2), for example on sugar beet molasses, can be achieved by Pseudomonas cepacia (Celik et al., 2005); or on cheese whey by Azotobacter vinelandii (Dhanasekar et al., 2001); or on molasses and whey by Rhizobium meliloti, Rhizobium viciae and Bradyrhyzobium japonicum (Mercan and Beyatli, 2005). It is also possible to use palm oil effluent for the production of PHAs. Japanese researchers described a two-stage process (Hassan et al., 1996). In the first stage, anaerobic treatment of the waste by palm oil sludge was carried out to obtain organic acids, particularly acetic and propionic acid. The acids were then converted into PHA by a phototrophic bacterium, Rhodobacter sphaer-oides, in the second stage.

The microbiological production of PD from glycerol is of great interest worldwide since its polycondensation with dicarbonic acids also results in

Table 9.2 Production of biopolymers

Substrate(s) Microorganism



Sugar beet molasses Cheese whey

Molasses and whey

Palm oil sludge Food wastes (whey, cane sugar)

Pseudomonas cepacia Azotobacter vinelandii Rhizobium meliloti, R. viciase and Bradyrhizobium japonicum Rhodobacter sphaeroides

Poly(3-hydroxybutyrate) Poly(3-hydroxybutyrate) Poly(3-hydroxybutyrate)

2005 Dhanasekar et al., 2001 Mercan and Beyatli, 2005

Poly(3-hydroxyalkanoate) Hassan et al.,


Poly(lactic acid) Datta et al.,


Table 9.3 Microbiological production of 1,3-propanediol





Glycerol Glycerol

Glycerol and dihydroacetone Glycerol


Clostridium butyricum Klebsiella pneumoniae

Klebsiella aerogenes

Citrobacter freundii

Clostridium pasteureanum


1,3-propanediol and

2,3-butanediol 1,3-propanediol



Papanikolaou et al., 2000 Biebl et al., 1998

Streekstra et al., 1987 Homann et al.,

1990 Heyndrickx et al., 1991

biodegradable polyesters. Two of the microorganisms used are Clostridium butyricum (Papanikolaou et al., 2000) and Klebsiella pneumoniae (Biebl et al., 1998) (Table 9.3). Beyond polyesters, poly(lactic acid) is another biopolymer with high potential in environmentally friendly packaging materials, it can be manufactured from lactic acid based on food wastes (Datta et al., 1995).

Microbial fat

Microbial fat provides an alternative to plant and animal fats, and oils, and can also be manufactured from food wastes. Bednarsky et al. (1986) used molasses supplemented with whey as a broth and fermentation was carried out by Candida curvata yeast species. The biomass contained triglycerides (11%) and proteins (23-34%).

204 Handbook of waste management and co-product recovery Miscellaneous compounds

Of the miscellaneous compounds, one of the most interesting groups is phytochemicals; for example, ellagic acid was synthesized by Lentidus edodes on cranberry pomace (Vattem and Shetty, 2003). A similarly remarkable and peculiar example is the production of bacterial cellulose by Acetobacter xylinum on untreated beet molasses in a loop airlift reactor (Bae and Shoda, 2005).

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