Beranda > Artikel Ilmiah > SIFAT PROOKSIDATIF FORTIFIKAN NaFeEDTA DAN Fe-SULFAT PADA KECAP HASIL FORTIFIKASI* (Oleh : Sri Naruki, Mary Astuti, Yustinus Marsono, dan Sri Raharjo**)

SIFAT PROOKSIDATIF FORTIFIKAN NaFeEDTA DAN Fe-SULFAT PADA KECAP HASIL FORTIFIKASI* (Oleh : Sri Naruki, Mary Astuti, Yustinus Marsono, dan Sri Raharjo**)

ABSTRAK

Telah dilakukan penelitian tentang pengaruh penambahan fortifikan NaFeEDTA dan FeSO4 pada kecap manis, terhadap angka peroksida dan angka TBARS komponen minyak, serta kadar senyawa karbonil komponen asam amino pada larutan kecap model. Sebagai kontrol, digunakan kecap tanpa fortifikasi. Dosis fortifikan adalah 266 mg Fe/L kecap. Pengaruh penambahan H2O2 terhadap peningkatan oksidasi juga dievaluasi. Berdasarkan data yang diperoleh dapat disimpulkan bahwa bila tidak ada penginduksi oksidasi, maka NaFeEDTA maupun FeSO4 tidak memicu oksidasi komponen minyak maupun asam amino.  Adanya penginduksi oksidasi, dalam hal ini H2O2, terbukti meningkatkan laju oksidasi, yang tercermin dengan adanya kenaikan angka peroksida, angka TBARS, dan kadar senyawa karbonil. Peningkatan laju oksidasi yang terjadi pada kecap hasil fortifikasi dengan NaFeEDTA tidak lebih besar daripada peningkatan pada kecap hasil fortifikasi menggunakan FeSO4 .

Kata kunci : fortifikasi, NaFeEDTA, angka peroksida, angka TBARS, senyawa karbonil

  

ABSTRACT

 Iron in iron-fortified soy sauce may has oxidative effect in both oil and amino acids or protein component. In order to evaluate the possible oxidative properties of both NaFeEDTA and FeSO4 fortificants, a study of oxidation in soy sauce fortified with the fortificants  was conducted. Level of fortificant was 266 mg Fe/L of soy sauce. The extent of oxidation was measured by the level of peroxide value and TBARS value of oil component of fortified soy sauce, as well as carbonyls content of soy sauce model. The effect of H2O2 addition on  level of oxidation was also evaluated. It was found that in soy sauce fortified with either NaFeEDTA or FeSO4,  the fortificants  did not promote oxidation of oil and  amino acids components. However, addition of H2O2 made peroxide value, TBARS value, and carbonyls content of fortified samples increased. 

 Keywords : fortification, NaFeEDTA, peroxide value, TBARS value, carbonyls content.

  

PENDAHULUAN

Defisiensi zat besi merupakan salah satu problema defisiensi zat gizi di dunia, termasuk Indonesia. Salah satu cara penanggulangan yang tampaknya menjanjikan adalah fortifikasi zat besi. Dari segi biaya, fortifikasi dianggap cukup efektif. Berbeda dengan pemberian tablet zat besi, fortifikasi ini dapat dengan mudah diterapkan  secara berkesinambungan pada populasi  yang menjadi target karena tambahan asupan zat besi dapat berlangsung tanpa disadari oleh konsumen (Anonim, 2001 dan  Fidler dkk., 2002).

Dalam fortifikasi, diperlukan adanya makanan pembawa, yang berperan sebagai media pembawa bagi fortifikan. Kecap kedelai manis dapat dipilih sebagai makanan pembawa karena produk tersebut digemari dan dikonsumsi secara meluas. Selain itu,  kecap memiliki warna coklat gelap dengan citarasa yang relatif kuat, sehingga perubahan warna dan citarasa yang mungkin timbul akibat fortifikasi, tidak menggangu kenampakan dan citarasa kecap. 

Fortifikasi zat besi menjadi tidak efektif bila diterapkan pada masyarakat dengan pola diet  yang  kaya dengan bahan nabati, seperti halnya yang terjadi di Indonesia. Hal ini disebabkan karena pada pangan nabati terdapat beberapa komponen yang dapat menghambat absorpsi zat besi oleh tubuh, antara lain senyawa fitat. Sebagai contoh,  absorpsi zat besi pada bahan nabati seperti jagung, bayam, dan gandum hanya mencapai 5%; jauh lebih kecil daripada absorpsi pada daging, yang dapat mencapai 10% (Reddy, dkk., 2000; Samman dkk., 2001; Lynch, 2002; dan Anonim, 2004).

Dari beberapa penelitian terakhir terungkap bahwa NaFeEDTA {sodium iron (Fe3+) ethylenediaminetetraacetic acid} dapat  meningkatkan bioavailabilitas zat besi fortifikan (Heimbach dkk., 2000 dan  Fidler dkk., 2002).  Hal ini disebabkan karena Fe pada EDTA tidak dapat diikat oleh senyawa fitat maupun senyawa penghambat lainnya, sehingga absorpsi zat besi menjadi tidak terganggu (Lynch, 2002). Fortifikan NaFeEDTA juga tidak menimbulkan perubahan warna dan citarasa, serta tidak mengakibatkan pengendapan selama penyimpanan produk (Fidler dkk., 2002). Studi pendahuluan menunjukkan bahwa kecap kedelai manis hasil fortifikasi menggunakan NaFeEDTA sebanyak 2.000 ppm tidak mengakibatkan perubahan sifat organoleptik dan tetap disukai panelis (Naruki, 2004, data belum dipublikasi).

Satu hal yang masih mendatangkan kontroversi adalah kekhawatiran bahwa  zat besi fortifikan dalam produk makanan hasil fortifikasi dapat menimbulkan efek oksidatif, baik terhadap komponen lemak/minyak maupun protein/asam amino. Oksidasi lemak/minyak dan protein dalam tubuh sering dikaitkan penuaan dini dan beberapa kondisi patologik seperti aterosklerosis, penyakit Alzheimer, dan artritis rematoid (Leeuwenburgh dkk., 1997; Berlett dan Stadtman, 1997; Shacter, 2000).

Dalam sistem pangan, oksidasi lemak/minyak akan menghasilkan peroksida, “thiobarbituric acid reactive substances” (disingkat TBARS), dan citarasa tengik. Zat besi diketahui dapat bertindak sebagai katalis dalam pembentukan TBARS (Raharjo, 2004). Secara umum, peroksida dan TBARS digunakan sebagai parameter tingkat kerusakan lemak/minyak akibat oksidasi. Dari segi organoleptik, citarasa tengik umumnya berperan sebagai faktor penentu dalam penerimaan konsumen terhadap produk (Gordon, 1990 dan  Santosa dkk., 1996).

Reaksi oksidasi terhadap protein akan menghasilkan senyawa karbonil. Semua jenis asam amino dapat terlibat dalam reaksi oksidasi. Sama halnya dengan oksidasi lemak, zat besi dapat bertindak sebagai katalis dalam oksidasi protein. Selain itu,  H2O2, sinar ultra violet, dan asap rokok diketahui dapat memacu pula terjadinya oksidasi (Yan dkk., 1997; Berlett dan Stadtman, 1997; Shacter, 2000).

Dalam diet, kecap berperan sebagai penyedap; bukan merupakan sumber minyak maupun protein. Hal ini disebabkan karena kecap mengandung minyak maupun protein dalam jumlah yang  relatif kecil dan dikonsumsi dalam jumlah yang relatif sedikit. Kadar minyak kecap umumnya lebih rendah dari 1%, sedangkan kadar protein berkisar antara 2 – 6% (Anonim, 1994).  Meskipun kadarnya rendah, oksidasi komponen minyak dan protein kecap menjadi penting karena peristiwa kimiawi tersebut dapat menghasilkan perubahan citarasa kecap yang tidak dikehendaki.    

Untuk mengetahui kemungkinan adanya sifat prooksidatif NaFeEDTA, telah dilakukan penelitian oksidasi pada kecap hasil fortifikasi dengan NaFeEDTA maupun Fe-sulfat. Tingkat oksidasi ditera melalui penentuan angka peroksida, angka TBARS, dan kadar senyawa karbonil. Selain itu, dipelajari pula pengaruh penambahan H2O2 terhadap tingkat oksidasi yang terjadi. 

 

METODA PENELITIAN

Bahan

Kecap manis merk tertentu, yang dibuat dari kedelai hitam (Glicine max L.) non-GMO, garam dapur (NaCl), gula kelapa, dan air,  tanpa penambahan bumbu, diperoleh dari toko swalayan di Yogyakarta. NaFeEDTA yang digunakan diperoleh dari Akzo Nobel CNF, Jakarta. Bahan kimia, antara lain Fe-sulfat (FeSO4.7H2O), Na-tiosulfat (Na2S2O3), thiobarbituric acid (TBA); 1,1,3,3-tetraetoxypropan (TEP); aseton; 2,4-dinitrofenylhidrazin (DNPH), dan asam amino, diperoleh dari MERCK.

Fortifikasi Kecap

Dalam penelitian ini digunakan tiga jenis sample, yaitu kecap tanpa fortifikasi (sebagai kontrol), kecap dengan 2.000 ppm fortifikan NaFeEDTA (setara 266 mg Fe/L

kecap), dan kecap dengan  FeSO4 (kadar 266 mg Fe/L kecap).   Untuk membuat kecap yang mengandung 2.000 ppm NaFeEDTA, 2.000 mg NaFeEDTA  dilarutkan dalam 20 mL aquades. Larutan tersebut ditambahkan ke dalam 1.000 mL kecap yang telah diketahui beratnya. Selanjutnya kecap  dipanaskan beberapa saat sampai dicapai berat semula. Fortifikasi kecap menggunakan Fe-SO4  dilakukan dengan cara yang sama. Untuk 1.000 mL kecap, diperlukan 1.301 mg FeSO4.7H2O, yang setara dengan 266 mg Fe.

Pembuatan Kecap Model

Dalam penelitian ini, kadar senyawa karbonil ditera secara spektrofotometrik. Warna coklat gelap pada kecap ternyata mengganggu peneraan absorbansi. Untuk mengatasi hal tersebut, sebagai sampel digunakan kecap model; yang mencerminkan kandungan asam amino kecap, namun tidak mengandung unsur pengganggu.

Sebelum pembuatan kecap model, pertama kali dilakukan analisis komposisi asam amino kecap. Berdasarkan data yang diperoleh, dibuat kecap model dengan jalan melarutkan beberapa asam amino dengan komposisi tertentu  (112 mg asam aspartat, 33 mg treonin, 42 mg serin, 505 mg asam glutamat,  55 mg glisin, 51 mg alanine, 39 mg valin, 31 mg isoleusin, 36 mg leusine, 22 mg tirosin, 25 mg fenilalanin, 11 mg histidin, 23 mg arginin, dan 40 mg prolin) ke dalam 100 mL H2O; sehingga diperoleh larutan model dengan komposisi asam amino yang sama dengan kecap. Fortifikasi kecap model dilakukan dengan cara yang sama seperti yang telah diterangkan sebelumnya.

Penentuan Angka Peroksida

Angka peroksida ditentukan menurut prosedur standar AOAC (1995). Mula-mula, minyak diekstrak dari 60g kecap hasil fortifikasi. Minyak hasil ekstraksi selanjutnya dilarutkan dalam 50 mL campuran asam asetat dan kloroform (3:2 v/v) dan digojog perlahan. Sebagai penginduksi oksidasi digunakan H2O2 (1 : 49 v/v) dengan variasi penambahan berturut-turut adalah  0, 125, 250, dan 500 mL. Penambahan H2O2 masing-masing dilakukan terhadap 10 mL larutan minyak tersebut. Selanjutnya dilakukan pula  penambahan 0,5 mL larutan KI jenuh dalam H2O. Kemudian sampel dibiarkan selama satu menit, untuk selanjutnya ditambah 30 mL H2O. Akhirnya sampel dititrasi dengan 0.01N Na-tiosulfat, dengan indikator amilum. AP dinyatakan sebagai  mili-equivalen peroksida/g minyak. 

Penentuan Angka TBARS

Angka TBARS ditentukan dengan menggunakan prosedur Tarladgis dkk. (1960). Sampel kecap hasil fortifikasi sebanyak 3 mL, dilarutkan dalam 100 mL H2O. Untuk menginduksi oksidasi, ke dalam larutan sampel dilakukan penambahan H2O2 (1 : 49 v/v). Variasi penambahan H2O2 yang dilakukan adalah 0, 500, dan 1000 mL.

TBARS diekstrak dengan cara distilasi pada 100 oC, setelah terlebih dahulu dilakukan penambahan 1,5 mL HCl 25% ke dalam larutan sampel. Proses ekstraksi dihentikan setelah diperoleh sekitar 75 mL distilat. Distilat ini selanjutnya ditambah H2O sampai volumenya mencapai 100 mL. Kemudian 1mL distilat tersebut dicampur dengan 1,5 mL H2O dan 2,5 mL larutan  TBA (0.02M TBA dalam asam asetat). Campuran selanjutnya dipanaskan pada suhu   100 oC selama 35 menit, menggunakan penangas air mendidih sehingga terbentuk senyawa kompleks berwarna merah jambu. Setelah didinginkan, terhadap campuran tersebut dilakukan pengukuran absorbansi secara spektrofotometri pada panjang gelombang 528nm. Sebagai standar, digunakan TEP. Angka TBARS dinyatakan dalam mmol MDA/g minyak.

Penentuan Kadar Senyawa Karbonil

Dalam analisa ini, sebagai penginduksi oksidasi digunakan H2O2, yang dibuat dengan melarutkan 1mL H2O2 dalam 499mL  H2O.  Kadar senyawa karbonil ditera secara spektrofotometrik dengan penggunaan DNPH sebagai bahan kimia yang spesifik terhadap karbonil, sebagaimana yang telah dilaporkan oleh Yan dkk. (1997).

Kecap model sebanyak 0,5 mL digunakan sebagai sampel analitik. Untuk menginduksi oksidasi, dilakukan penambahan H2O2 ke dalam sampel. Variasi penambahan H2O2 berturut-turut adalah 0, 50, 100, dan 150 mL; disusul dengan penambahan H2O, berturut-turut sebanyak 500, 450, 400, dan 350 mL, sehingga dicapai volume akhir masing-masing sebesar 1,0mL. Kemudian dilakukan penambahan 1,0 mL  DNPH  dan satu tetes HCl 37 %. Campuran tersebut dipanaskan pada suhu 50 oC selama 30 menit, menggunakan penangas air; disusul dengan pendinginan dengan air mengali dan penambahan 8.0mL KOH (1,0 N). Setelah penggojogan, absorbansi campuran diukur dengan spektrofotometer pada panjang gelombang 480 nm. Sebagai standar digunakan aseton. Kadar senyawa karbonil dinyatakan sebagai  mg aseton/mL larutan kecap model.

 

HASIL DAN PEMBAHASAN

 

Pengaruh NaFeEDTA terhadap Angka Peroksida

            Evaluasi tentang pengaruh fortifikan zat besi terhadap angka peroksida komponen minyak yang diekstrak dari   kecap hasil fortifikasi,   menghasilkan data   yang dapat dilihat pada Tabel 1.

Tabel 1. Angka peroksida komponen minyak kecap hasil fortifikasi, dengan atau tanpa penambahan H2O2

 

Angka peroksida

(mili-equivalen peroksida/g minyak)

               Penambahan H2O2 (mL)

 

Fortifikan

 

0

 

125

 

250

 

500

Kontrol

0,06g

2,64e

5,03c

9,38a

NaFeEDTA

0,02i

1,92f

3,72d

7,57b

Fe-sulfat

0,05h

2,04f

3,68d

6,98b

Catatan:   Huruf yang berbeda menunjukkan adanya perbedaan nyata dengan a 0,05

           

Data pada tabel tersebut menunjukkan bahwa fortifikasi kecap dengan zat besi, baik dalam bentuk NaFeEDTA maupun Fe-sulfat, terbukti menghasilkan angka peroksida yang lebih rendah daripada kontrol. Hal ini disebabkan karena senyawa peroksida (ROOH) merupakan produk antara (“intermediate products”) dari rangkaian reaksi oksidasi minyak. Adanya zat besi akan memicu laju reaksi lanjutan senyawa peroksida tersebut, dengan produk akhir berupa senyawa aldehid dan keton. Laju degradasi senyawa peroksida yang berlangsung lebih cepat tersebut akan menurunkan akumulasi senyawa peroksida yang ada.  Pada kontrol, laju reaksi degradasi senyawa peroksida  berlangsung lebih lambat sehingga akumulasi senyawa peroksida akan relatif lebih banyak. Penambahan H2O2 mengakibatkan peningkatan angka peroksida, baik pada sampel  hasil fortifikasi dengan NaFeEDTA maupun Fe-sulfat. Keduanya menghasilkan peningkatan angka peroksida yang sama besar.

 

Pengaruh  NaFeEDTA terhadap Angka TBARS

               Tabel 2 menunjukkan pengaruh jenis fortifikan dan penambahan H2O2  terhadap angka  TBARS. Bila tidak ada “inducer” H2O2, fortifikasi kecap menggunakan NaFeEDTA maupun Fe-sulfat terbukti tidak menaikkan angka TBARS. Jadi dapat dinyatakan bahwa dalam kondisi yang terlindung dari pemicu oksidasi, maka fortifikasi kecap dengan 2.000 ppm NaFeEDTA tidak menimbulkan pengaruh yang merugikan terhadap komponen minyak kecap.

Tabel 2. Angka TBARS komponen minyak kecap hasil fortifikasi, dengan atau tanpa penambahan H2O 

 

Angka TBARS (mg malonaldehid/g minyak)

        Penambahan H2O2 (mL)

 

Fortifikan

 

0,0

 

0,5

 

1,0

Kontrol

8,12e

56,97d

114,90c

NaFeEDTA

7,78e

132,04bc

160,25b

Fe-sulfat

6,73e

112,03c

273,78a

Catatan: Huruf yang berbeda menunjukkan adanya perbedaan nyata dengan a 0,05

             

            Data yang diperoleh juga menunjukkan bahwa penambahan penginduksi H2O2 dapat meningkatkan angka TBARS, baik pada sampel kecap hasil fortifikasi dengan NaFeEDTA maupun Fe-sulfat.  Fortifikan   NaFeEDTA  mengakibatkan  peningkatan   angka TBARS yang tidak lebih besar daripada fortifikan Fe-sulfat. Ini berarti bahwa sifat prooksidatif  NaFeEDTA  relatif lebih kecil daripada sifat prooksidatif Fe-sulfat.  

Pengaruh NaFeEDTA terhadap Kadar Senyawa Karbonil

            Data tentang pengaruh jenis fortifikan dan penambahan H2O2  terhadap kadar senyawa karbonil larutan kecap model dapat dilihat pada Tabel 3. Data yang diperoleh menunjukkan bahwa bila tidak  ada   penginduksi H2O2,  maka   NaFeEDTA maupun Fe-sulfat tidak meningkatkan laju oksidasi asam amino, sehingga  menghasilkan  kadar senyawa karbonil yang sama dengan kontrol. Jadi dapat dinyatakan bahwa dalam kondisi yang terlindung dari pemicu oksidasi, maka fortifikasi kecap dengan 2.000 ppm NaFeEDTA tidak mengakibatkan pengaruh yang merugikan terhadap komponen asam amino.

Table 3. Kadar senyawa karbonil kecap model, dengan atau tanpa penambahan H2O2

 

Kadar senyawa karbonil

(mg aseton/mL larutan model)

                  Penambahan H2O2 (mL)

 

 

Fortifikan

 

0

 

50

 

100

 

150

Kontrol

0,54d

6,39d

7,72d

8,87d

NaFeEDTA

3,51d

58,46c

82,52b

108,86a

Fe-sulfat

14,53d

64,14c

87,29b

110,81a

Catatan:  Huruf yang berbeda menunjukkan adanya perbedaan nyata dengan a 0,05

 

            Pada kecap model yang hanya mengandung asam amino, penambahan  H2Odapat meningkatkan produksi senyawa karbonil, sebagai produk oksidasi asam amino. Produksi senyawa karbonil tersebut semakin meningkat dengan semakin banyaknya H2Oyang ditambahkan. Data yang diperoleh juga menunjukkan bahwa fortifikan   NaFeEDTA maupun Fe-sulfat  mengakibatkan  peningkatan   kadar senyawa karbonil yang sama besar. Ini membuktikan bahwa sifat prooksidatif  NaFeEDTA  tidak lebih besar daripada Fe-sulfat. Dengan demikian, penggunaan NaFeEDTA sebagai fortifikan diharapkan dapat meningkatkan bioavailabilitas zat besi, namun tidak mendatangkan pengaruh yang merugikan terhadap komponen minyak maupun asam amino/protein kecap.

 

KESIMPULAN

Berdasarkan data yang diperoleh dapat disimpulkan bahwa keberadaan NaFeEDTA maupun Fe-sulfat  dalam kecap hasil fortifikasi (266 mg Fe/L kecap), tidak memicu oksidasi komponen minyak maupun asam amino, jika tidak ada pemicu oksidasi. Namun demikian,  H2O2, dapat  meningkatkan laju oksidasi, yang tercermin dengan adanya kenaikan angka peroksida, angka TBARS, dan kadar senyawa karbonil.

 

UCAPAN TERIMA KASIH

 Peneliti mengucapkan banyak terima kasih kepada  PT. Unilever Indonesia Tbk., Jakarta, Indonesia, atas bantuan yang diberikan, baik berupa bantuan finansial maupun fortifikan NaFeEDTA. Selain itu, ucapan terima kasih juga disampaikan kepada QUE Project, Jurusan Teknologi Pangan dan Hasil Pertanian, Fakultas Teknologi Pertanian Universitas Gadjah Mada Yogyakarta, atas bantuan finansial yang diberikan demi kelancaran penelitian ini.

 

DAFTAR  PUSTAKA

Anonim (1994). Mutu dan Cara Uji Kecap. Standar Nasional Indonesia. Dewan Standardisasi Nasional.

Anonim (2001). Food Fortification Development in Indonesia. Food Fortification Commission of Indonesia, UNICEF, Center for Food and Nutrition Policy Studies – Bogor Agricultural University, and Food Communication Forum of Indonesia.

Anonim (2004). Dietary Supplement Fact Sheet : Iron. Office of Dietary Supplement. Warren G. Magnuson Clinical Centre. National Institute of Health. USA.

AOAC (1995). Official Methods of Analysis of AOAC International. Sixteenth Edition, Volume I. AOAC International, Maryland, USA.

Berlett, B.S. dan Stadtman, E.R. (1997). Protein oxidation in aging, disease, and oxidative stress. Journal of Biological Chemistry  272 : 20313-20316.

Fidler, M, Davidsson, L., Walczyk, T. dan Hurrell, R.F. (2002). Iron bioavailability from iron fortified fish sauce and soy sauce. Eidgenossische Tehnische Hochshule. Zurich.

Gordon, M.H. (1990). The mechanism of antioxidant action in vitro. Dalam :  Hudson, B.J.F. (ed.). Food Antioxidants, hal. 1-18.  Elsevier Applied Science, London dan New York.

Heimbach, J., Rieth, S., Mohamedshah, F., Slesinki, R., Samuel-Fernando,P., Sheehan, T., Dickman, R. dan Borzelleca, J. (2000). Safety assesment of iron EDTA [sodium iron (Fe3+) ethylenediaminetetraacetic acid] : summary of toxicological, fortification, and exposure data. Food and Chemical Toxicology  38 : 99-111.

Leeuwenburgh, C., Rasmussen, J.E., Hsu, F.F.,  Mueller, D.M., Pennathur, S. dan   Heinecke, J.W. (1997). Mass spectrometric quantification of marker for protein oxidation by tyrosyl radical, copper, and hydroxyl radical in low density lipoprotein isolated from human atherosclerosis plaques. Journal of Biological Chemistry 272 : 3520-3526.

Lynch, S. (2002). Food iron absorption and its importance for the design of food fortification strategies. Nutrition Review 60 : S3-S6.

Naruki, S. (2004). Fortifikasi Kecap dengan NaFeEDTA. Data belum dipublikasi.

Raharjo, S. (2004). Kerusakan Oksidatif pada Makanan. Pusat Studi Pangan dan Gizi. Universitas Gadjah Mada. Jogjakarta, Indonesia.

Reddy, M.B., Hurrel, R.F. dan Cook, J.D. (2000). Estimation of nonheme-iron bioavailability from meal composition. American Journal of  Clinical Nutrition  71 : 937-943.

Santosa, U., Kubo, K., Ota, T., Tadokoro, T. dan Maekawa, A (1996). Antioxidative effect of coconut (Cocos nucifera L.) water extract on TBARS value in lever of rat fed fish oil diet. Indonesian Food and Nutrition Progress  3 : 42-50.

Samman, S., Sandstrom, B., Toft, M.B., Bukhave, K., Jensen, M., Sorensen, S.S. dan  Hansen, M. (2001). Green tea or rosemary extract added to foods reduces nonheme-iron absorption. American Journal of Clinical Nutrition 73 : 607-612.

Shacter, E. (2000). Quantification and significance of protein oxidation in biological samples. Drug Metabolism Review 32 : 307-326.

Tarladgis, B.G., Watts, B.M. dan Younathan, M.T. (1960). A distilation method for the quantitative determination of malonaldehyde in rancid foods. Journal of American Oil Chemistry Society  37 : 44.

Yan, L.J., Lodge, J.K., Traber, M.G., Matsugo, S. dan Packer, L. ( 1997). Comparison between copper-mediated and hypochlorite-mediated modifications of human low density lipoproteins evaluated by protein carbonyl formation. Journal of Lipid Research 38 : 992-1001.

 

*)  Hasil penelitian ini sudah dipublikasikan di Majalah Ilmiah AGRITECH –  Fakultas Teknologi Pertanian – UGM.

**) Penulis adalah Staff pengajar di Jurusan Teknologi Pangan dan Hasil Pertanian ,  Fakultas Teknologi Pertanian, Universitas Gadjah Mada, Yogyakarta.

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