10 Chymosin in Cheese Making 243Rennet paste, a rich source of lipolytic enzymes,
has been a major factor in the flavor development of
ripened Italian cheeses such as Provolone and Ro-
mano. Conceivably, 2100 years ago, Romans applied
rennet paste to cheese milks to develop a typical
“picante” flavor in Romano and related cheeses, for
even then calf rennet was used to set their cheese
milks. In more modern times many countries, in-
cluding the United States, applied calf rennet paste
to induce flavor, largely for Italian ripened cheeses.
Farntiam et al. (1964) successfully produced,
from goats, dried preparations of a pregastric-oral
nature, also rich in lipolytic enzymes. They applied
them to milk for ripened Italian cheese with excel-
lent results. Since then, lipase powders, of various
character and strength, have largely replaced rennet
paste.
CHYMOSINPRODUCTION BYGENETIC
TECHNOLOGY
Genetic technology has been used for the commer-
cial production of a 100% pure chymosin product
from microbes. This type of chymosin is often
called fermentation-produced chymosin. Uchiyama
et al. (1980) and Nishimori et al. (1981) published
early on the subject, and in 1990 Pfizer, Inc.,
received U.S. Food and Drug Administration
approval to market a genetically transformed prod-
uct, Chy-Max, with GRAS stature (Duxbury 1990).
Other brands using other microorganisms have also
been approved. Table 10.1 lists some examples of
such products.
The microbes used for this type of rennet include
nonpathogenic microorganisms Escherichia coliK-
12, Kluyveromyces marxianus var. lactis and
Aspergillus nigervar. awamori. Prochymosin genes
obtained from young calves are transferred through
DNA plasmid intervention into microbial cells.
Fermentation follows to produce prochymosin, cell
destruction, activation of the prochymosin to chy-
mosin, and harvesting/producing large yields of
pure, 100% chymosin. This product, transferred
from an animal, is considered a plant product, as
microbes are of the plant kingdom. Thus, they are
acceptable to various religious groups.
These products have been widely studied to eval-
uate their impact on the quality and yield of cheese.
An initial concern was that because of the absence
of pepsin from these types of rennets, proper cheese
flavor might not develop. Most studies have general-
ly concluded that there are no significant differences
in flavor, texture, composition, and yield compared
with calf rennet controls (Banks 1992, Barbano and
Rasmussen 1992, Biner et al. 1989, Green et al.
1985, Hicks et al. 1988, IDF 1992, Ortiz de Apodaca
et al. 1994).
When these products were first introduced, it was
expected that a majority of cheese makers would
convert to these types of rennets because they are
virtually identical to traditional calf rennet and
much cheaper. Except for some areas, this has not
generally happened because of some concerns to-
wards genetically modified products. Further, some
protected cheeses, such as those under French AOC
regulations or Italian DOP regulations, require theTable 10.1.Examples of Some New Chymosin ProductsProduct Name Company and Type
AmericanPure Sanofi Bio-Industries
Calf rennet purified via ion exchange
Chy-max Pfizer (Now Chr. Hansen)
Fermentation—Using Escherichia coliK-12
Chymogen Chr. Hansen
Fermentation—Using Aspergillus nigervar. awamori
ChymoStar Rhône-Poulenc (Now Danisco)
Fermentation—Using Aspergillus nigervar. awamori
Maxiren Gist-Brocades
Fermentation—Using Kluyveromyces marxianusvar. lactis
Novorena Novo Nordisk
(Marzyme GM) Fermentation—using Aspergillus oryzae
aNovoren is not chymosin: it is the coagulating enzyme of Mucor mieheicloned into As. oryzae. The other products listed
above are all 100% chymosin.