SUMMARY

Enzyme Modified Cheese (EMC) Flavour Generation with Aspergillus niger Lipase and Lactobacillus casei Aminopeptidase and Application of EMC on

Accelerating Gouda Cheese Ripening

By: Abdul Manab, Manik E. Sawitri, Khothibul Umam Al A.

Animal Product Technology Department Faculty of Animal Husbandry, Brawijaya University

This research was conducted from March until November 2008 in the Laboratory of

Animal Product Technology - Faculty of Animal Husbandry, Laboratory of Agricultural Product Technology - Faculty of Agriculture Technology, Central Laboratory of Food Science and Technology - Brawijaya University and Laboratory of Molecular Biology - Faculty of Mathematic and Science, Brawijaya University. Laboratory of Matematic and Science Faculty of Mathematic and Science - Gadjah Mada University.

The objective of this research was gouda cheese ripening acceleration using EMC which emulsified with lechitine. The objective of this research was gouda cheese ripening acceleration using EMC which emulsified with lechitine.

Materials used were whey protein, beef tallow sapi, curd gouda cheese, L. casei, A. niger, A. niger Lipase, L. casei protease, maltodextrin, arabic gum. Equipment used were analitic balance (Ohaus BC Series dan Mettler Instruments, Perancis), pH meter (Hanna Instruments, Perancis), dan spektrofotometer UV (Thermospectronic Genesysttm 10 UV), water bathr, vortex-mixer, autoclave, incubator, pipetman, GC, GCMS, HPLC (Hitachi), CLSM.

The research method on A. niger Lipase production with beef tallow addition (0, 4 and 8%) and L. casei protease production with whey protein addition (0, 4 and 8%) using Completely Randomised Design. The measured variables were cell production, enzyme production and activity. The research method on EMC production with lipase addition (0, 0,3 and 0,6%) was Completely Randomised Design. The measured variables were fat content, Free Fatty Acid, Amino acid Profile, Fatty Acid Profile and EMC Flavour. The research method on EMC encapsulation with Maltodextrin and Arabic Gum (0, 3 and 6% respectively) were Completely Randomised Design. The measured variables were Lipase and protease activity, water content, flavour retention and microencapsulation retention. The research method on EMC Stabilisation during storage was Completely Randomised Design. The measured variables were Lipase and protease activity, Free Fatty Acid, peroxide, flavour, amino acid and fatty acid profile.

Result of variance analysis indicated that treatment of beef tallow addition did not gave a significant effect (P>0.05) on A. Niger production, gave very significant effect (P<0,01) on lipase production and activity. Treatment of whey protein addition did not gave significant effect (P>0,05) on L. casei protease activity, however gave very significant effect (P<0,01) on protease production. Addition of A. niger lipase gave significant (P<0,05) on water content,and EMC fat content, gave very significant effect (P<0,01) on EMC FFA, however didnot gave significant effect (P>0,05) on EMC soluble protein. Arabic gum addition as encapsulation agent gave very significant effect (P<0,01) on water content, activity of EMC powder lipase and protease, however did not gave significan (P>,0,05) on EMC powder microencapsulation efficiency. Maltodextrin addition as encapsulation agent gave very significant effect (P<0,01) on

water content, activity of EMC powder lipase and protease, however did not gave significan (P>,0,05) on EMC powder microencapsulation efficiency. Storage of EMC powder increase lipase activity FFA and peroxide, change flavour, amino acid and fatty acid profile.

Research showed that lechitine did not gave significant effect (P>0,05) emulsion stability of gouda cheese. Using of EMC did not gave significant effect (P>0,05) on water content, lipid content, TPC, lightness and yellowness of gouda cheese, gave highly significant effect on (P<0,01) on protease, lipase activity, LAB, FFA and texture gouda cheese and gave significant effect (P<0,05) on redness gouda cheese. Ripening time on EMC gave highly significant effect (P< 0,01) on water content, lipid content, protein content, pH, protease activity, lipase activity, LAB, TPC, FFA, Lighness, redness, yellowness and texture of gouda cheese. EMC 2% gave more compact microstructure and better potein-lipid distribution; more branched amino acid, more aromatic amino acid and more methionine; more short chain fatty acid; more flavor produced among ripening.

Based on research concluded that beef tallow addition (8%) increase lipase production and activity, whey powder addition (8%) increase protease production and activity. Addition of lipase 0,6% produce better EMC characteristic. Using Maltodextrin 6% and arabic gum 3% produce better EMC powder (lipase and protease activity, efficiency microencapsulation). Storage EMC powder during 15 and 30 days increase lipase and protease activity, peroxide and FFA. It was suggested So produce EMC powder using 0,3% lipase and encapsulant agent (maltodextrin 6% or arabic gum 3%), storage EMC powder no more than 15 days on room temperature. It was suggested to research about EMC powder emulsification and application EMC powder on accelerating gouda cheese ripening.

It concluded that: lechitine improve emulsion stability EMC powder in gouda cheese EMC; 2% EMC improve chemical composition (water, lipid, protein), decrease pH, LAB, TPC; increase FFA, lipase and protease activity, texture; improve lightness, redness and yellowness, more compact microstructure. EMC 2% accelaraed gouda cheese ripening. It suggested using EMC powder 2% on acceleration gouda cheese ripening.

RINGKASAN

Pembangkitan Flavour Enzyme Modified Cheese (EMC) dengan Lipase Aspergillus niger dan Aminopeptidase Lactobacillus casei Serta Aplikasinya

terhadap Percepatan Pemeraman Keju Gouda

Abdul Manab, Manik E. Sawitri, Khotibul Umam A.

Program Studi Teknologi Hasil Ternak Fakultas Peternakan Universitas Brawijaya ■ Penelitian ini dilakukan mulai bulan Maret 2008 sampai Nopember 2009 di Laboratorium

Teknologi Hasil Ternak Fakultas peternakan, Laboratorium Sentral Ilmu Hayati Universitas Brawijaya. Laboratorium Biologi molekuler Fakultas MIPA Universitas Brawijaya, dan Laboratorium MIPA universitas Gadjah Mada.

Penelitian Tahun I ini bertujuan peningkatan produksi enzim lipase melalui penambahan lemak s pada media pertumbuhan A. Niger, Peningkatan produksi enzim protease melalui penambahan whey bubuk pada media pertumbuhan L. Casei, Pembangkitan flavour EMC menggunakan lipase A. niger dan protease L. asei, Enkapsulasi enzim dan flavour EMC menggunakan maltodekstrin dan gum arab selama proses pengeringan, Stabilitas enzim lipase dan aminopeptidase serta retensi flavour EMC bubuk selama penyimpanan. Penelitian Tahun II ditujukan untuk percepatan waktu pemeraman keju gouda dengan penambahan EMC yang diemulsifikasi menggunakan lesitin.

Materi yang digunakan dalam penelitian ini adalah protein whey, lemak sapi, curd keju gouda, L. Casei, A. niger, Lipase A. niger, protease L. casei, maltodekstri, gum arab. Peralatan yang digunakan di antaranya adalah timbangan analitik {Ohaus BC Series dan Mettler Instruments, Perancis), pH meter (Hanna InstrumentsvPerancis), dan spektrofotometer UV (Thermospectronic Genesysttm 10 UV), penangas air, vortex-mixer, autoclave, inkubator, pipetman, GC, GCMS, HPLC (Hitachi), Chromameter dan CLSM.

Penelitian Tahun I produksi lipase dari A. niger dengan perlakuan lemak sapi (0, 4 dan 8%) dan protease L. casei dengan perlakuan protein whey (0, 4 dan 8%) menggunakan Rancangan Acak Lengkap. Variabel diuji pada penelitian ini adalah produksi sel, produksi dan aktifitas enzim. Produksi EMC dengan perlakuan lipase A. niger (0, 0,3 dan 0,6%) menggunakan rancangan acak lengkap, variabel yang diamati: kadar lemak, protein terlarut, kadar FFA, profil asam lemak, profil asam amino dan flavour EMC. Enkapsulasi EMC menggunakan maltodekstrin dan gum arab dengan perlakuan masing-masing (0, 3 dan 6%) dengan suhu pengering semprot 130°C menggunakan rancangan acak kelompok dengan variabel yang diamati kadar air, aktifitas lipase serta protease, efisiensi enkapsulasi dan retensi flavour. Penyimpanan EMC pada suhu ruang dengan waktu penyimpanan 1, 15 dan 30 hari dengan variabel yang diamati adalah aktifitas lipase dan protease, kadar FFA, Peroksida, profil asam lemak, asam amino dan stabilitas flavor. Penelitian Tahun II penggunaan lesitin keju gouda dengan mikrostruktur yang lebih kompak dengan sebaran lemak dan protein yanglebih merata; asam amino lebih banyak mengandung asam amino rantai cabang, mengandung sulfur dan aromatic; lebih banyak mengandung asam lemak rantai pendek serta mengandung flavor yang lebih banyak lebih cepat pembentukannya.

Berdasarkan hasil penelitian Tahun I disimpulkan bahwa Penggunaan lemak sapi sebanyak 8% menghasilkan produksi lipase dengan aktifita yang lebih tinggi, dan protein whey

sebanyak 8% menghasilkan protease lipase dengan aktifitas yang lebih tinggi. Penggunaan lipase sebanyak 0,6% menghasilkan karakteristik EMC yang lebih baik. Penggunaan Maltodekstrin sebanyak 6% dan gum arab sebanyak 3% menghasilkan EMC bubuk dengan efisisensi enkapsulasi, aktifitas lipase dan protease yang lebih baik. Penyimpanan EMC bubuk pada suhu ruang selama 15 dan 30 hari berakibat pada peningkatan aktifitas lipase dan protease dan perubahan peroksida, FFA, Flavour, profil asam lemak dan asam amino. Disarankan Produksi EMC bubuk menggunakan lipase 0,3% dan dibutuhkan enkapsulan berupa maltodekstrin 6% atau gum arab 3% dan penyimpanan pada suhu ruang sebaiknya tidak melibihi 15 hari. Perlu kajian lebih lanjut tentang emulsifikasi EMC bubuk pada keju dan aplikasi EMC bubuk untuk percepatan pematangan keju gouda.

Kesimpulan dari penelitian tahun ke-2 ini adalah: Penambahan lesitin dapat meningkatkan stabilitas emulsi EMC bubuk dalam keju gouda, Penambahan EMC bubuk sebanyak 2% berpengaruh pada komposisi kimia keju (kadar air, kadar lemak, kadar protein), menurunkan pH, jumlah BAL dan TPC, meningkatkan FFA, aktifitas protease dan lipase, membentuk struktur keju menjadi lebih kompak, memperbaiki warna Kecerahan (L), kemerahan (a) dan kekuningan (b) serta menghasilkan senyawa flavor lebih banyak dan lebih cepat, Penambahan EMC bubuk sebanyak 2% dapat mempercepat pematangan keju gouda. Disarankan untuk Percepatan pematangan keju bisa menggunakan penambahan EMC bubuk sebanyak 2%, Diperlukan penelitian lebih lanjut tentang karakterisasi peptida, MAG, DAG dan senyawa flavour yang potensial bagi kesehatafi.

DAFTAR PUSTAKA

Alinkolis, JJ. 1979. Candy Technology. The AVI Publishing Company. Westport Connecticut. Anonymous. 1998. Microencapsulation. http://www.wsu.edu/~gmhyde /433 web pages/dryinff-

web-pages98/spray-dry/microencapsulation.htm. Tanggal akses: 04-09-2006. Anonymous, 2001. Spesifikasi By-product Bogasari. PT. ISM Bogasari Flour Mills. Surabaya. Anonymous, 2006. The Use of Enzyme for Dairy Flavour Enhancement (including Enzyme

Modified Cheese). Biocatalysts limited Cefn Coed Pare Nantgarw CF 15 7QQ Wales UK. WWW.biocatalysts.com

Anonymous. 2007. Aspergillus niger. http://permimalang.wordpress.com/ 2007/12.htm. Diakses Tanggal 30 Juni 2008.

Arora G. and B. H. Lee, 1992. Purification and characterization of aminopeptidase from L. casei ssp casei LLG. J. Daiiry Scie 75:700-710.

Baehaki, A., M.T. Suhartono, N.S. Palupi dan T. Nurhayati, 2008. Purifikasi dan Karakterisasi Protease dari Bakteri Patogen Pseudomonas aeruginosa. Jurnal.Teknol. dan Industri Pangan Vol. XIX No. 1: 80-86.

Balassa, L.L., and G.P. Fanger. 1971. Microencapsulation in The Food Industry. CRC Crit. Rev. Food Technol. 2:245.

Bhandary, B.R., and D'Arcy. 1996. Microencapsulation of Flavour Compound. Technical Review in Food. Journal Australia Food. 48(2):547-551.

Buckle, K.A., R.A. Edward., G.H. Fleet., and M. Wootton. 1987. Ilmu Pangan: Penerjemah Hari Purnomo dan Adiono. Ul-Press. Jakarta.

Burrington, K.J., 2002. The whey to nutrition. A technical resources for dairy manufacture. Wisconsin.

Choi H, L. Laleye, G.F. Amantea and R.E. Simard, 1996. Production of aminopeptidase from skim whey permeate medium by L. casei ssp casei. J. Dairy scie 79:956-063.

Cholette H., and R.C. McKellar, 1990. Influence of pH on the properties of I. helveticus aminopeptidase. J. Dairy scie 73:2278-2286.

Clark, J.R., 1992. Whey: Composition, properties, processing and technology. Volume 4. Hui, Y.H. (ed) Marcell Dekker Inc. New York.

D'Annibale A., G.G. Shermanni, F. Federici and M. Petruccioli, 2005. Olive-mill wastewaters: a promising substrate for microbial lipase production. Bioresource tech.

Dhanang, P. 2008. Pertumbuhan Sel Bakteri. http://www. Multiply.Global.curr_reply_to.id/coretan%20dan%20catatan____%20dha nang%20-%20pertumbuhan%20sel%20bakteri_files/36.htm. Diakses Tanggal 23 Oktober

2006. Dinata, D.I. 2004. Mutasi Kapang Monascus sp. dengan Etil Metana Sulfonat dan Analisis Kadar

Sitrinin Hasil Fermentasi Cair Galur Induk dan Mutannya. Thesis. Institut Teknologi Bandung. Bandung

Drgalic, I., L. Tratnik, and R. Bozanic, 2005. Growth and Survival of Probiotic Bacteria in Reconstituted Whey. Le Lait 85: 171-179.

Durham, D.R., 1990. The unique stability of Vibrio proteolyticus neutral protease under alkaline conditions afford a selective step for purification and use in amino acid-coupling reaction. Appl. Environ. Microbiol. 56: 2277-2281.

Dziezak, J.D. 1988. Microencapsulation and Encapsulated Ingredient. Food Technol. 42:135.

Fachruddin, L. 1998. Memilih dan Memanfaatkan Bahan Tambahan Makanan. Trubus Agriwidya. Ungaran.

Fallony, G., J. C. Armas, J.C.D. Mendoza and J.L.M. Hernandez, 2006. Production of Extracellular lipase from Aspergillus niger by Solid-State Fermentation. Food Technol. Biotechnol 44 (2) 235-240.

Fawzya, Y.N., 2002. Karakterisasi Protease Ekstraseluler dari Isolat Bakteri Asal Ikan Hiu Atas (Carcharhinus limbatus). Tesis IPB. Bogor.

Fennema, O.R. 1985. Food science. Marcel and Dekker, Inc. New York. Fox, P.F and P.L.H. McSweeney. 1991. Rennets: Their Role in Milk Coagulating and Cheese

Ripening. In Law, B.A: Microbiology and Biochemistry of Cheese and Fermented Milk. 2nd Ed. Chapman and Hall. London.

Fox, D.F., T.P. Guinee, T.M. Logan and P.L.H. McSweeney. 2000. Fundamentals of Cheese Science. An Aspen Publication. Gaithersburg. Maryland.

Fressner, W.D. 2000. Biocatalysis from Discovery to Application. Springer Verlag. Berlin. Galloway, J. H and R. J. M. Grawford. 1985. Cheese Fermentation. In Daulay, D. 1991.

Buku/Monograf Fermentasi Keju. PAU Pangan dan Gizi. IPB. Bogor. Iyer, M., T. Richardson, C.H. Amundson and R.C. Tripp. 1967. Major Free Fatty Acid in Gouda

Cheese. J. Dairy Sci. 50(3): 385-392. Ha E., and Zemel M.B., 2003. Functional Properties of Whey, Whey Components, and Essential

Amino Acids: Mechanism Underlying Health Benefits for Active People. J. Nutr. Biochem. 14:251-258.

Hengenbart, S. 1993. Encapsulated Ingredients Keep Problem Covered. Food Product Design. April, 21-25.

Hill, C.G., S. Ghannouchi and H.S. Garcia, 2001. Lypolisis of butter oil by immobilized lamb pregastric esterase: I. Uniresponse kinetics-pH and Temperature Effects. J. Dairy science, 84:1034-1043.

Jay, J.M. 1992. Modern Food Microbiology. 4th edition. Chapman and Hall. New York. Kamini N.R., J.G.S. Mala and R. Puvanakrishnan, 1998. Lipase production from Aspergillus

niger by solid state fermentation using gingelly oil cake. Process Biochem 33 (5):505-511.

Kamzolova S.V., I.G. Morgunov, A. Aurich, O.A. Perevoznikova, N.V. Shishkanova, U. Stottmeister and T.V. Finogenova, 2005. Lipase secretion and citric acid production in Yarrowia lipolytica yeast grown on animal and vegetable fat. Food Technol Biotechnol 43 (2) 113-122.

Ketaren, S. 2008. Pengantar Teknologi Minyak dan Lemak Pangan. UI Press. Jakarta. Khosrowshasi, A., A. Madadlau, M. E. Z. Mousavi, and Z. E. Djomeh. 2006. Monitoring the

chemical and textural changes during rippening of Iranian white cheese made with different concentrations of starter. J. Dairy Sci. 89(5):3318-3325.

Kieronczyk A., S. Skeie, T. Langsrud and M Yvon, 2003. Cooperation between Lactococcus lactis and Nonstarter Lactococci in the formation of cheese aroma from amino acids. Applied and Environmental Microbiology, Feb. 2003 p. 734-739.

Kilcawley K.N., M.G. Wilkilson, and P.F. Fox, 2001. A Survey of Lipolytic and Glycolitic End-Products in Commercial Cheddar Enzyme-Modified Cheese. J. Dairy Science (84)66-73.

Kombong, H. 2004. Evaluasi Daya Hidrolitik Enzim Glukoamilase dari Filtrat Kultur Aspergillus niger. J. Ilmu Dasar. 5 (1): 16-20.

Kravtchenko, T.P. 1998. The Use of Acacia Gum as a Source of Soluble Dietary Fiber. In Mosiihey, S.H. 2003. Influence of Different Capsule Materials on The Physiological Properties of Microencapsulated Lactobacillus acidophilus. Inaugural - Dessertation. Departement of Food Technology. University of Bonn. German.

Krochta, J.M., E.A. Baldwin., and M.O. Nisperos. 1994. Edible Coatings and Films to Improve Food Quality. Technomic Publishing Company Inc. New York.

Lee, K.T, T.A. Foglia dan M.J. Oh. 2002. Food Chemistry and Toxicology. Medium Long Medium and Medium Long Long Acylglycerol from Beef Tallow and Caprylic Acid. J. Food Sci. Vol. 67(3): 1-6

Longo, M. A. and M.A. Sanroman, 2006. Production of Food Aroma Compunds: Microbial and Enzymatic Methodologies. Food Tecnol. Biotechnol 44(3):335-353.

Mahadik, N.D., K.B. Bastawde, U.S. Puntambekar, J.M. Khire and D. V. Gokhale, 2002. Production of acidic lipase by a mutant of Aspergillus niger NCIM 1207 in submerged fermentation. Process Biochem 39 (12):2031-2034

Marshal, T., 1995. Standart Dairy Methods. John Willey and Sons. New York. Master, K. 1985. Spray Drying Handbook. Fourth Edition. Longman Scientific.

Mathews, C. K., K. E. Van Holde dan K.G. Ahern. 2000. Biochemistry, 3rd edition. Addison Wesley Longman Inc. San Francisco.

McNamee B.F., E.D. O'Riordan and M. O'Sullivan, 2001. Effect of Partial replacement of gum Arabic with carbohydrates on its microencapsulation properties. J. Agric Food Chem 49 (7):3385-3388.

Me Williams and Margaret. 1997. Food Experimental Prespective. Third Edition. Prentice Hall Inc. New Jersey.

Mosilhey, S.H. 2003. Influence of Different Capsule Materials On The Physiological Properties of Microencapsulated Lactobacillus acidophilus.

Inaugural - Dessertation. Departement of Food Technology. University of Bonn. German. Naiola, E. dan N. Widhyastuti, 2002. Isolasi, Seleksi dan Optimasi Produksi Protease dari

Beberapa Isolat bakteri. Berita Biologi. 16 (3): 467-473. __________________________, 2007. Semi Purifikasi dan Karakterisasi Enzim Protease Bacillus sp. Berk. Penel. Hayati 13: 51-56. Nurhidayati, P. 2003. Pengaruh Konsentrasi Enzim Papain dan Suhu Fermentasi Terhadap

Kualitas Keju Cottage. KAPPA ISSN 1411-4046. Vol. 4, No. 1: 13-17. Nurika, I. 2000. Pengaruh Konsentrasi Dekstrin dan Suhu Inlet Spray Dryer Terhadap Stabilitas

Warna Bubuk Pewarna dari Ekstrak Angkak. Jumal Teknologi Pertanian. Vol. 1 No. 2 : 15-23.

Pomeranz, Y. dan Cairman, 1973. Industrial Uses of cereals. A.A.C.C. St Louis. Missouri. Putranto, R.A., D. Santoso, T. Panji, Suharyanto dan A. Budiani. 2006. Karakterisasi Gen

Penyandi Lipase dari Kapang Rhizopus oryzae dan Absidia corymbifera. J. Menara Perkebunan. 74 (1): 23-32.

Rahayu, K. 1986. Isolasi dan Pengujian Aktivitas Enzim. PAU Pangan dan Gizi Universitas Gadjah Mada. Yogyakarta.

Ranhotra, G.S., Gelroth, J.A., Glase, B.K. and Reddy, P.V., 1994. Nutritional Profile of a Fraction from Air-Classified Bran Obtained from a hard red wheat. J. Cereal chem. 7 (4):321-324.

Roosdiana, A., H. Kartikaningsih, Suharjono, R. Peranginangin dan Murdinah, 2003. Isolasi dan Karakterisasi Bacillus sp Penghasil Protease dari Kulit Ikan Kakap Merah {Lutjanus sanguineus). Jurnal Ilmu-ilmu Hayati. Vol. 15 No. 2:139-145.

Rosenberg, M., I.J. Kopelman and Y. Talmon, 1990. Factors affecting retention in spray drying microencapsulation of volatile materials. J. Agric. Food chem. 38:1288-1294.

Scott, R.M. 1986. Cheese Making Practice. 2nd Ed. Elsevier Applied Sci. Publ. London. Singh T.K. M.A.M Drake and K.R. Cadwallader, 2003. Flavor of Cheddar cheese: a chemical

and sensory perspective. Food science and food safety vol 2:139-162. Shoutgate, O.A.T. 1991. Determination of Food Carbohydrates. Second Edition. Elsevier

Applied Science. London and New York. Singh, T.K., M.A.M Drake and K.R. Cadwallader. 2003. Flavour of Cheddar Cheese: a

Chemical and Sensory Perspective. Food Science and Food Safety. Vol 2:139-162. Soottitantawat A., H. Yoshii, T. Furuta, M. Ohkawara and P. Linko, 2003. Microencapsulation

by spray drying: influence of emulsion size on the retention of volatile compounds, J. Fodd Science 68(7): 2256-2262.

Stanbury, P. F. and A. Whitaker. 1984. Principles of Fermentation Technology. Pergamon Press Ltd. Oxford

Srbinovska, S., T. Cizbanovski and V. Dzabirski. 2001. Dynamics of salt diffusion and yield of three types of goat's milk cheese. J. Dairy Sci. 51 (1): 15-26.

Tarakci, Z and E. Kucukoner. 2006. Changes on phisicochemical, lipolysis, and proteolysis of vacuum packed Turkish Kashar cheese during rippening. J. Central Europ. Agric. 7(3):459-464.

Tsakalidou E., I. Dalezios , M. Georgalaki and G. Kalantzopoulos, 1993. A Comparative study: aminopeptidase activities from L. bulgaricus and S. thermophilus. J. Dairy Scie 76:2145-2151.

Tungjaroenchai, W., C.H. White., W.E. Holmes and M.A. Drake. 2004. Influence of Adjunct Cultures on Volatile Free Fatty Acids in Reduced Fat Edam Cheeses. J. Dairy Sci. 87:3224-3234.

Winarno, F.G. 1984. Enzim Pangan. PT. Gramedia Pustaka Utama. Jakarta. Walstra, P. A., T. J. Geurts., A. Noomen., A. Jellema and M. A. J. S. Van Boekel. 1993 .Dairy

Technology Principles of Milk Properties and Processes. Dutch-Type Varieties Departement of Food Science. Wageningen Agricultural University. Wageningen. Netherlands. Marcel Dekker, Inc. New York

Widodo, 2003. Bioteknologi Industri Susu. Lacticia Press. Yogyakarta. Winarno, F.G. 1984. Enzim Pangan. PT. Gramedia Pustaka Utama. Jakarta. Winarno, F. G. 1993. Pangan, Gizi, Teknologi, dan Konsumen. PT Gramedia Pustaka Utama.

Jakarta. --------- 2002. Kimia Pangan dan Gizi. PT Gramedia Pustaka Utama. Jakarta. Winarno, F. G. 2004. Kimia Pangan dan Gizi. PT Gramedia Pustaka Utama. Jakarta. Yvon M., S. Thirouin, L. Rijnen, D. Fromenteher and J.C. Gripon, 1997. An Aminotransaminase

from L. lactis initiates conversion of amino acids to - cheese flavour compounds. App Environ Microb 63(2):414-419.