K. PNEUMONIAE VE P. MIRABILIS TESPİTİ İÇİN TOZ ANTOSİYANİN İÇERİKLİ-ÜREAZ TESTİNİN RASYONEL HAZIRLANMASI

Öz

Bu çalışmada, üreaz-pozitif bakteriler olan Klebsiella pneumoniae ATCC 54475 ve Proteus mirabilis ATCC 25933'ün hızlı, seçici, hassas tespiti için Brassica oleracea var. Capitata (kırmızılahana) lliyofilize ekstresi ilize ekstresi (RCE) içeren kolorimetrik ortam geliştirilmiştir. Ana bileşen kırmızı lahana, pH'a bağlı renk değişiminden sorumlu antosiyanin gruplarıdır. Üre testi, 0.01 M sodium fosfat tampon ve sodium azit içinde hazırlanmış belirli antosiyanin, üre konsantrasyonu içerir. Testteki üre, K. pneumoniae ve P. mirabilis'ten salınan üreaz enzimi ile hidrolize edilir ve sırasıyla test ortamını alkalin yapan, antosiyaninin proton kaybetmesine neden olan ve pembeden yeşile renk değişimini indükleyen amonyum (NH3) molekülleri üretir. Ayrıca Escherichia coli ATCC 25922, üreaz negatif kontrol bakterisi olarak kullanıldı. K. pneumoniae ve P. mirabilis'in tespiti, antosiyanin konsantrasyonu ve inkübasyon süresinin bir fonksiyonu olarak incelenmiştir. Antosiyanin, hidroklorikasit (HCl) ilavesi nedeniyle sıvı formdaki serbest protonun girişimini atmak için üreaz testinde liyofilizetoz formunda kullanıldı. Görsel sonuçların kantitatif teyiti için dijital görüntü analizi olan Delta E (∆E) ölçümleri ile sağlanmıştır. Sonuçlar, K. pneumoniae ve P. mirabilis için sırasıyla 150 ve 210 dakika inkübasyonda pembeden yeşile renk değişiminin gözlendiğini göstermektedir. Burada, antosiyanin içeren üreaz testini başarıyla hazırlayarak sonuçların çıplak göz ve dijital görüntü analizi ile K. pneumoniae ve P. mirabilis'in kolorimetrik tespitini gerçekleştirdik.

Anahtar Kelimeler

• antosiyanin
• dijital görüntü analizi
• hızlı üreaz testi
• üreaz pozitif bakteri

RATIONAL PREPARATION OF POWDER ANTHOCYANIN INCORPORATED UREASE TEST FOR DETECTION OF K. PNEUMONIAE AND P. MIRABILIS

Öz

In this present study, we developed Brassica oleracea var. Capitata (red cabbage) extract (RCE) powder- incorporated colorimetric media for rapid, selective, sensitive detection of urease-postive bacteria, Klebsiella pneumoniae ATCC 54475 and Proteus mirabilis ATCC 25933. The major component of RCE is anthocyanin groups which are in charge of pH dependent color change. The ureas test contains certain concetration of anthocyanin, urea prepated in 0.01 M sodium phosphate buffer and sodium azide. The urea in the test was hydrolyzed by urease enzyme released from K. pneumoniae and P.mirabilis and produce ammonium (NH3) molecules which sequentially made the test media alkaline, caused deprotonation of anthocyanin and induced the color change from pink to green. Also, Escherichia coli ATCC 25922 was used a urease-negative control bacteria. The detection of K. pneumoniae and P. mirabilis was examined as a function of anthocyanin concentration and incubation time. The anthocyanin was used lyophilized powder form in the urease test to discard the in interference of free proton in liquid form owing to addition of hydrochloric acid (HCl). We also performed the digital image analysis, Delta E (∆E) for quantitative confirmation of the visual results. It was observed that the results showed that the color change from pink to green in 150 and 210 minutes of incubaiton for K. pneumoniae and P. mirabilis, respectively. Here in, we successfully prepared anthocyanin-incorporated urease test and perform colorimetric detection of K. pneumoniae and P. mirabilis with naked eyes and digital image analysis.

Anahtar Kelimeler

• rapid urease test
• Anthocyanin
• digital image analysis
• urease-postive bacteria

Kaynakça

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