Dairy Chemistry And Biochemistry

410 DAIRY CHEMISTRY AND BIOCHEMISTRY

Depending on the depth of information required, proteolysis in cheese is

assessed by a wide range of techniques. Electrophoresis, usually urea-

PAGE, is particularly appropriate for monitoring primary proteolysis, i.e.

proteolysis of the caseins and the resulting large polypeptides. Quantifying

the formation of peptides and amino acids soluble in water, at pH 4.6, in

TCA, ethanol or phosphotungstic acid, or the measurement of free amino

groups by reaction with ninhydrin, o-phthaldialdehyde, trinitrobenzene or

fluorescarnine, is suitable for monitoring secondary proteolysis. Reversed

phase HPLC is especially useful for fingerprinting the small peptide profile

in cheese and is now widely used. High-performance ion-exchange or size

exclusion chromatography are also effective but are less widely used.

Proteolysis has not yet been fully characterized in any cheese variety but

considerable progress has been made for Cheddar and, as far as is known,

generally similar results apply to other low-cook, internal bacterially

ripened cheeses (e.g. Dutch types). Proteolysis in Cheddar will be sum-

marized as an example of these types of cheese.

Urea-PAGE shows that a,,-casein is completely hydrolysed in Cheddar

within 3-4 months (Figure 10.19). It is hydrolysed by chymosin, initially at

Phe,,-Phe,, and later at Leu,,,-Lys,,,, and to a lesser extent at Phe,,-

Gly,,, Leu,,-Lys,, and Leu,,,-Glu, Although p-casein in solution is

readily hydrolysed by chymosin, in cheese /3-casein is very resistant to

chymosin but is hydrolysed slowly (c. 50% at 6 months) by plasmin at

Lys,,-Lys,,, Lys,,,-His/Gln,,, and Lys,,,-Glu,,,, producing yl, yz- and

y3-caseins, respectively, and the corresponding proteose-peptones (PP5,

PP8 slow and PP8 fast; Chapter 4). Chymosin and, to lesser extent, plasmin

C I2 3 4 5 6 7 8 9 1011121314

Figure 10.19 Urea-polyacrylamide gel electrophoretograms of Cheddar cheese after ripening
for 0, 1, 2, 3, 4, 6, 8, 10, 12, 14, 16, 18 or 20weeks (lanes 1-14); C, sodium caseinate. (Supplied
by S. Mooney.)