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KAJ) Kurdistan Academicians Journal, 2004, 3(1) Part A ( A 2004 ) 1 (3

Short Note The flora of fermentation and changes of acidity during the production of Kurdish food Doina
Huner H. Arif* and Bahrouz M. A. Al jaff
Department of Biology, College of Science, University of Sulaimani, Kurdistan Region, Iraq.

Abstract
To evaluate the fermentation flora in the fresh phase of the Kurdish food, Doina, microbiological analysis conducted within fermentation period (96 h). Lactic acid bacteria increased from 2.39×105 to 8.65 ×107 cfu/g accompanied by decreasing in pH from 4.88 to 4.81 which increased later to 4.87. Acidity started with 0.171%, increased and decreased then to 0.225%. Yeasts increased from 1.7 to 5.65×107 cfu/g. Proteolytic bacteria decreased from 4.6×107 to 6.1×106 cfu/g within 48 h, increased then to reach 7.15×107 cfu/g at 72 h, declined later to 8×106 cfu/g. Lipolytic bacteria increased from 3×107 to 7.1×108 cfu/g at 72h then declined to 2.075×108 cfu/g. It was appeared that the fermentation of Doina is of lactic acid type.

Key words:- fermentation, bacteria, yeast, Doina. Introductio
Fermentation process is one of the most ancient forms of food preservation methods in the world, it is cheap and energy efficient means of preserving perishable raw materials which increase their shelf life. In Kurdistan, there are several types of food preserved by fermentation of which is Doina, a fermentative product of a mixture of Do; buttermilk (the Kurdish synonym of the agitated milk; churned milk), bulgur (crushed wheat) and salt. There were no available reports on fermentative bacteria, spoilage flora as well as, the pathogenic and enterotoxin producer bacteria that may contaminate them. However, there are some studies on related foods which revealed lactic acid bacteria the main fermentative flora included Nuruk, Kishk, Rabadi or Rabdi and Gergoush [1,2,3,4]. Doina made from concentrated Do mixed with bulgur and salt, the mixture then placed in an anaerobic conditions (underground) for 4-7 days. After ripening, the product taken out, mould as balls and dried by exposuring to sunlight. There is no any previous literature about this type of food while related foods were subjected to some investigations, for example, Kishk is an Egyptian (and most of Arabian countries) fermented product prepare from parboil wheat and milk which finally forms into small balls and sundried [2]. Microorganisms which are responsible for fermentation of Kishk include Lactobaillus plantarum, Lactobaillus brevis, Lactobaillus casei, Bacillus subtilis and yeasts [5,6]. This study was aimed to evaluate the fermentation flora and the change of acidity in the fresh phase of Doina.

Materials and Methods
Microbiological analyses: They conducted for the detection of lactic acid, proteolytic, lipolytic bacteria and yeasts as total colony forming units (cfu) per gm. Samples were prepared by

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KAJ) Kurdistan Academicians Journal, 2004, 3(1) Part A ( A 2004 ) 1 (3

Results
Initiated fermentation was with high number of lactic acid bacteria 2.39×105 cfu/g and yeast 1.7×107 cfu/g (Figure 1). The lactic acid bacteria increased to reach a maximum count within 72h 8.95×107 cfu/g before they

4.8 4.78 4.76 0 24 47 72 96 0.05 0

Time (hours)
pH Total acidity

Figure(2): The change of acidity(%) and pH during the 96h of Doina fermentation

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Total Acidit (%) y

pH

homogenizing 5g of fresh Doina with 45ml sterile distilled water by blending, then serial 10fold dilutions were made. Detection of lactic acid bacteria (LAB): aliquots of 0.1ml of required dilutions of fresh Doina samples (along the fermentation period) were spread on Rogosa agar plates [7], and incubated in an anaerobic jar for 3 days at 32˚C. After incubation, the colonies considered lactic acid bacteria were 0.5-2 mm diameter, small greyish-white, flat or raised, smooth, rough or intermediate. Detection of proteolytic bacteria: aliquots of 0.1ml of required dilutions of fresh Doina as a bove were spread on skim milk agar (20% of 10% skim milk solution in nutrient agar) and incubated at 32˚C for 48h, the appearance of clear zone around colonies indicated the proteolytic bacteria. Detection of lipolytic bacteria: aliquots of 0.1 ml of required lipid agar (30 ml of lipoidal emulsion made up of 1ml tween 80 added to 400ml warm distilled water and 100ml of olive oil emulsified in a blender until the oil being homogenized and autoclaved, were added to one litre of nutrient agar) and incubated at 32˚C for 48 h, the medium appeared opaque but lipolytic colonies were surrounded by a zone of clear medium. Detection of yeasts: aliquots of 0.1ml of required dilutions of fresh Doina were spread on sabouraud dextrose agar (oxoid) and incubated at 32˚C for 48 h. pH determination [8]: pH measurements were determined by homogenizing 10g of fresh Doina with 20ml sterile distilled water using a blender. The pH of the homogenate was then measured with a pHmeter (LABQUIP, England). Total acidity (%) determination: Titratable acidity as lactic acid was determined by titration of fresh Doina samples against down-ward NaOH volume [9].

declined slightly to reach 8.65×107 cfu/g. The same happened with yeasts which increased to reach 3.5×107 cfu/g within 24h. This number remains stable for 3 days before it increased at the end stage of fermentation to reach 5.65×107 cfu/g (Figure 1).
cfu/gm(107 )
100 10 1

0.1 0 0 24 48 72 96

(Time (hours
Lactic acid Bacteria Lipolytic Bacteria
X

Yeasts Proteolytic Bacteria

The lipolytic bacteria increased from 3× 107cfu/g to a maximum level at 72 h fermentation 7.1× 108 cfu/g prior to decline sharply at 96 h of fermentation to become 2.075×108 cfu/g (Figure 1). The proteolytic bacteria declined at the first stages of fermentation from 4.6×107 cfu/g at zero time to 6.1×106 cfu/g within 48 hours of fermentation. They then increased to a maximum level at 72 h fermentation to 7.15×107 cfu/g and declined again sharply after 96 h of fermentation to 8×106 cfu/g the end time of fermentation (Figure 1). The total acidity (%) was increased from 0.171 at the zero time to 0.225 after 96h. This acidity led to decrease pH 4.9 0.25 value from 4.88 at zero time to 4.81 within 48 h 4.88 0.2 but the relative stability of lactic acid bacteria 4.86 0.15 led to increase the pH value to 4.87 at 96h 4.84 fermentation stage (Figure 2). 4.82 0.1

Figure(1): The change in bacterial and yeast number during 96h fermentation process of Doina

KAJ) Kurdistan Academicians Journal, 2004, 3(1) Part A ( A 2004 ) 1 (3

Discussion
In Doina, the initiation of fermentation step starts by addition of agitated milk which is a lactic acid fermentation product serves as a starter of fermentation. In Gergoush (a related Sudanian food) which ingredients are legume, boiled milk and wheat flour, three genera of bacteria dominated which included lactic acid bacteria (LAB), Bacillus spp. and Clostridium spp. [4]. Nearly the same observations were with the Korean food Nuruk [1] and the Indian fermented food product Rabadi [3]. The similar food Egyptian Kishk was showed to contain Lactobacillus plantarum, L.brevis, L.casei as well as Bacillus subtilis and yeasts [5,6] but there are no reports about the lactic acid bacterial counts within the fermentation process. The

similar and identical food found in the Arabian regions of Iraq called Chishch but with no studies carried on. So, Doina is considered as a lactic acid fermentation product. Because of addition of agitated milk in Doina which has somewhat high lipid content, the lipolytic bacteria increased. Kishk, reported to contain 23.5% protein content [5,6]. So, Doina is expected to have high protein content also. The initial fall of proteolytic bacteria may be due to the dominance of lactic acid bacteria and yeasts, which compete them, sugars are metabolized firstly by lactic acid organisms then lipolytic and proteolytic organisms increased (Figures 1 & 2). It was concluded that lactic acid bacteria and yeasts are the fermentative flora during Doina fermentation. The lactic acid, lipolytic, proteolytic bacteria and yeasts should be identified at the species level to study the succesion of microflora during fermentation and even till spoilage.

References
1. Kim, C. J. Microbiological and enzymological studies on Takju brewing. J. Korean Agricul. Chem. Soc., 1968, 60, 69-99 (cited by FAO, 1999 Fermented cereals, a global perspective. FAO agriculture services bulletin No. 138). 2. Morcos, S. R.; Hegazi, S. M. & Ell-Damhoughy, S. I. T. Fermented foods common in Egypt I. Nutritive value of Kishk. Journal of the Science of Food and Agriculture. 1973a, 24, 1153-1156 (cited by FAO, 1999 Fermented cereals, a global perspective. FAO agriculture services bulletin No. 138 ). 3. Sankaran, R. Fermented foods of the India subcontinent. In: Wood, B. J. B. (ed.) Microbiology of Fermented Foods, vol. 2, 2nd edn., Thompson science, 1998, 753-789. 4. Sherfi, S. A. & Hamad, S. H. Microbiological and biochemical studies on Gergoush fermentation. Int. J. Food Microbiol., 2001, 47(3), 247-252 (Absract). 5. Beuchat, L. R. Indigenous fermented foods. In: Reed, G. (ed.) Food and Feed production with Microorganisms. Biotechnology, 1983, vol. 5, Verlag Chemie, Weinheim 477 (cited by FAO, 1999 Fermented cereals, a global perspective. FAO agriculture services bulletin No. 138 ). 6. Odunfa, S. A. African Fermented Foods. In: Wood, B. J. B. (ed.) Microbiology of Fermented Foods, 1985,vol. 2, Elsevier Applied Science Publishers, London & New York (cited by FAO, 1999 Fermented cereals, a global perspective. FAO agriculture services bulletin. 138 ). 7. Rogosa, M.; Mitchell, J. A. & Wiseman, R. F. A selective Medium for the Isolation of Oral and Fecal Lactobacilli. J. Bacteriol., 1951, 62(1), 132-133. 8. Little, C. L. & Knochel, S. Growth and survival of Yersinia enterocolitica, Salmonella and Bacillus cereus in Brie stored at 4, 8 and 20oC. Int. J. Food Micribiol.,1994, 24 137-145. 9. Brocklehurst, T. F. & Lund, B. M. Microbiological changes in cottage cheese varieties during storage at +7 C. Food Microbiol., 1985, 2, 207-233.

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‫( ‪KAJ) Kurdistan Academicians Journal, 2004, 3(1) Part A‬‬ ‫‪A‬‬ ‫4002‬ ‫3( 1 )‬

‫فلؤراي هةويَنكاري و طؤرِاني ترشيي لة كاتي بةرهةمهيَناني خؤراكي‬ ‫كوردي, دؤينة‬
‫هونةر هيوا عارف* و بةهرؤز مةحمود ئةمين جاف‬ ‫بةشي بايؤلؤجي/ كؤليجي زانست/ زانكؤي سليَماني/ هةريَمي كوردستان, عيراق.‬
‫بة مةبةستي خةملَندني فلؤراي هةويَنكاري لة خؤراكي كوردي (دؤينة) لة قؤناغي‬ ‫تةرِيدا,شيكاري مايكرؤبايؤلؤجي بة ئةنجام طةيةنرا لة ماوةي ثرؤسةي هةويَنكاريدا كة ماوةكةي‬ ‫69 كاتذميَرة. ذمارةي بةكترياي ترشي لكتيك لة 93.2×015 يةكةي دروستكردني نيشتةبةند (ي‬ ‫دن)/طرام بةرز بؤتةوة بؤ 56.8×017 ي دن/طرام لةطةل دابةزينيَك لة بةهاي تواني هايدرؤجيني‬ ‫‪ pH‬لة 88.4 بؤ 18.4 بؤ ئةوةي جاريَكي تر بةرز بيَتةوة بؤ 78.4.ريَذةي ترشيي لة 171.0% دةستي‬ ‫ثيَكردوة و بةرزبؤتةوة و دواي ئةوة جاريَكي تر دابةزيوة بؤ 522.0%. ذمارةي هةويَنةكان بةرز‬ ‫بؤتةوة لة 7.1×017 ي دن/طرام بؤ 56.5×017 ي دن/طرام. ذمارةي بةكترياي ثرؤتين شيكةرةكان‬ ‫دابةزيوة لة 6.4×017 ي دن/طرام بؤ 1.6×016 ي دن/طرام لة ماوةي 84 كاتذميَردا ثاشان بةرز‬ ‫بؤتةوة بؤ 51.7×017 ي دن/طرام لة ماوةي 27 كاذيَر و تا لة كؤتاييدا دابةزيوة بؤ 8×016 ي‬ ‫8‬ ‫دن/طرام. ذمارةي بةكترياي رؤن شيكةرةكان بةرز بؤتةوة لة 3×017 ي دن/طرام بؤ 1.7×01 ي‬ ‫دن/طرام لة ماوةي 27 كاذيَر و جاريَكي تر دابةزيوة بؤ 570.2×018 ي دن/طرام. دةركةوت كة‬ ‫ثرؤسةي هةويَنكاري دؤينة لة جؤري هةويَنكاريةكاني بةكترياي ترشي لكتيكة.‬

‫ثوختة‬

‫فلورا التخمير و تغير الحموضة عند انتاج الغذاء الكردي 0دوينه‬
‫هونه ر هيوا عارف * و بهروز محمود أمين الجاف‬ ‫قسم علوم الحياة /كلية العلوم/جامعة السليمانية/إقليم كردستان -العراق.‬

‫الخلصة‬
‫لغرض تقييم فلورا التخمير في الطور الطري للغذاء الكردي 0دوينه 0أجريت التحليلت‬ ‫الميكروبايولوجية ضمن فترة التخمير والتي أمدها 69 ساعة. إزداد العدد الكلي لبكتريا حامض‬ ‫اللبنيك من 93.2×015 إلى 56.8×017 وحدة تكوين مستعمرة (وت م)/غرام مصحوبة بهبوط في‬ ‫قيمة الس الهيدروجيني ‪ pH‬من 88.4 إلى 18.4 والتي إزداد لحقا َ إلى 78.4. بدأت الحموضة عند‬ ‫7‬ ‫171.0% و إزداد لتتنخفض ثانية إلى 522.0%. إزداد العدد الكلي للخمائر من 7.1×017 إلى 56.5×‬ ‫01 وت م/غرام. إنخفضت البكتريا الحالة للبروتين من 6.4×017 إلى 1.6×016 وت م/غرام عند 84‬ ‫ساعة فيما إزدادت إلى 51.7×017 وت م/غرام و عادت لتهبط أخيرا َ إلى 8×016 وت م/غرام في‬ ‫نهاية عملية التخمير. إزدادت البكتريا الحالة للدهون من 3×017 إلى 1.7×018 وت م/عرام عند 27‬ ‫ساعة و من ثم هبطت إلى 570.2×018 وت م/غرام. لقد بدا تخمير الدوينة علىإنه من نوع‬ ‫تخميرات حامض اللبنيك.‬

‫‪Received‬‬

‫وةرطيرا لة 3002/21/41 وثةسةندكرا لة 4002/3/11دا.‬ ‫4002/3/11 ‪.14/12/2003Accaptead‬‬

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KAJ) Kurdistan Academicians Journal, 2004, 3(1) Part A ( A 2004 ) 1 (3

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