Whey is an abundant byproduct of the dairy industry and can present a disposal problem due to its high BOO of approximately 40,000 mg
r1•
lt has been estimated that 47 % of the whey produced worldwide remains unused (Wessingeret al. , 1 990). While for smaller processing plants the return of whey to agriculture
may be economically favourable, for larger plants processes such as drying, protein recovery, whey fermentation, and lactose production may be more advantageous (Sienkiewicz and Riedel, 1 990). As the production of whey is constantly increasing, established processes for whey utilization m ust be improved and new solutions for processing the excess sought.
2.5.2 TYPES OF WHEY
Whey is the fluid obtained by separating the coagulum from milk, cream , and/or skim milk (Short, 1 978). Different types of whey are produced with 'sweet whey', having a pH value g reater than 5.5, derived from the manufacture of cheese or rennet casein, and 'acid whey', with a pH value less than or equal to 5.0, being obtained from cottage cheese, lactic acid casein, or mineral acid casein. Whey contains lactose, serum proteins, mineral salts, rennet, lactic o r mineral acid, and water. lt is a dilute material containing about 6 %(w/v) total solids, of which the major constituent (> 70 %) is lactose (Sienkiewicz and Riedel, 1 990). The serum protei ns are usually recovered as a whey protein concentrate (WPC). They show a range of functional properties when soluble and have a high nutritive value in both the soluble and insoluble form (Short, 1 978). Table 2.5 gives typical
compositions of deproteinated sweet and acid wheys.
Table 2.5 Composition of deproteinated sweet and acid wheys of various sources. (Adapted from Short, 1 978).
Parameter total solids (g kg"1) total nitrogen (g kg-1) non-protein nitrogen (g kg-1) ash (g kg-1) lactose (g kg-1)
lactic acid lactate (g kg"1)
ashffS (%) lactose!fS (%) pH Cheddar cheese 57.0 0.26 0.24 5.0 49.0 1.4 8.8 86.0 5.5 2.5.3 UTILIZATION OF WHEY Whey source
Cottage Lactic Sulphuric cheese casein casein
58.0 56.8 56.4 0.33 0.64 0.37 0.30 0.46 0.32 5.6 5.7 7.9 43.0 44.8 46.0 4.4 3.4 9.7 10.0 14.0 74.0 79.0 82.0 5.0 5.0 5.0
Table 2.6 details the volumes of whey (processed and non-processed) produced in two recent seasons in New Zealand. A breakdown of the usage of whey for these two seasons is also shown in Table 2.6.
In New Zealand, the major products fro m whey processing are whey protein concentrate (WPC), lactose a,nd lactalbumin (Barnett, 1 993) and examples of end-uses are products such as infant formulae, meat sausages, biscuits, chocolate drinks and cheese spreads. Sienkiewicz and Riedel (1 990) also list many other properties and uses.
Sienkiewicz and Riedel (1 990) noted a number of options for the use of deproteinated whey in fermentation processes, with examples shown i n Table 2.7. Whilst many processes are proposed only a limited number have been commercialized, and of these only ethanol is produced in New Zealand. The first
Table 2.6 Summary of quantities of whey processed in New Zealand over the 1 991 /92 and 1 992/93 seasons (Jim Barnett, 1 993).
1 991/92 Season 1 992/93 Season
Volume % of whey Volume % of whey (m3 yr-1) produced (m3 yr-1) produced
Whey produced 3023981 31 1 8770
Whey processed 1 82 1 874 60.3 2289 1 05 73.4
Non processed whey 201 233 39.7 829664 26.6
Processed whey % of processed % of processed
whey whey WPC 42.0 38.3 Whey powder 5.6 5 . 1 Whey cheese 1 .0 0.5 Lactose 20.9 29.7 Ethanol 5.8 (26.8t 4.7 Infant formula 7.1 7.9 Lactalbumin 1 7.6 1 3 .9
Non processed whey % of non % of non
processed whey processed whey
Fertilizer 46.5 46.5
Biological treatment 0.1 0.2
Animal 8.1 1 1 .7
River/sea 1 7.7 3.7
Irrigation 27.6 37.9
a Value in brackets is ethanol produced from the deproteinated whey of WPC and lactalbumin.
Table 2.7 Examples of fermentation products from whey or whey permeate.
Fermentation products
solvents biogas
polysaccharides
organic acids and their derivatives enzymes others Examples ethanol, butanol methane xanthan
amino acids, lactic, acetic, and citric acids �-galactosidase
caretenoids, flavour compounds, fats and oils, starter cultures, and whey yeast
distillery was commissioned by the New Zealand Co-operative Dairy Co. Ltd (NZCDC) at Reporoa in 1 980. The four distilleries now in operation at Reporoa, Tirau, Clandeboye, and Edgecumbe, supply the New Zealand market for both industrial and potable ethanol with an increasing proportion also exported, primarily to Japan. Mawson (1 987) recently reviewed the whey to ethanol i ndustry of New Zealand.
2.5.4 USE OF RECOMBINANT ORGANISMS FOR WHEY FERMENTATION
There has been interest in using S. cerevisiae to ferment whey, however this
yeast does not ferment lactose since it lacks both lactose permease and � galactosidase activity (Chen and Chiger, 1 985). To overcome these problems a number of techniques have been used, including the coimmobilizatio n of S.
cerevisiae cells and �-galactosidase to ferment whey permeate o r deproteinated
whey (Marwaha and Kennedy, 1 984; Hahn-Hagerdal, 1 985; Roukas and Lazarides, 1 991 ) . However, disadvantages include the relatively high cost of � galactosidase, and the diauxic g rowth of S. cerevisiae on glucose and g alactose
(Roukas and Lazarides, 1 991 ).
An alternative to techniques such as coi mmobilization is to use recombina nt D NA technology to construct �-galactosidase-positive strains of S.cerevisiae.
Sreekrishna and Dickson (1 985) have cloned and expressed both the l actose permease and �-galactosidase of K. lactis in S. cerevisiae, and Porro et al.
(1 991 ; 1 992) and Compagno et al. (1 993) have expressed an E. coli �
galactosidase in S. cerevisiae. ·The former recombinant strain grew on lactose as
a carbon source, but the rate of growth was low. The transformant produced by Porro et al. (1 991 ) had a high growth yield in rich lactose media and also s howed
high production of ethanol. Farahnak et al. (1 986) have constructed fusion strains
but such strains remained sensitive to inhibition by lactose. Alternatively, whey could be used as a substrata for production of other heterologous p rotei ns. Harlander (1 989) has noted that, for economic reasons, whey will probably be used to produce heterologous proteins using yeasts. To date no literature exists
for the production of a heterologous protein in K. lactis using whey as the
medium.
2.5.4 SU MMARY
Significant developments in the field of whey utilization have taken place over the last few decades. Whey in liquid form has limited use and so the concentration, drying and fermentation of whey, as well as the isolation of individual whey constituents have become increasingly important. As dairy processing plants in New Zealand become more centralized and therefore larger, the problem of disposing of excess non-processed whey also increases. lt is therefore desirable to find higher-returning products to increase the amount of whey processed. With recent advances in molecular biology techniques the yeast K. lactis could be
engineered to produce additional products to ethanol from whey and a new process developed.
Ideally, as a result of increased whey utilization, there would be less pollution, increased sales in presently manufactured whey products, and markets for improved and new whey products.
CHAPTER 3
MATE RIALS AND METHODS