Publication: Flavonoidlerin ve metal komplekslerinin yüksek basınçlı sıvı kromatografisi yöntemiyle yan yana belirlenmesi
Abstract
Bu çalışmada, Kuersetinin alüminyum(III) ve bakır(II) ile oluşturduğu komplekslerin kararlılık sabitleri Calvin-Bjerrum ve Irwing-Rossotti yöntemleri kullanılarak potansiyometrik yoldan tayin edildi. Kuersetinin protonlanma sabitleri potansiyometrik yöntemle logK1 = 11.15, logK2 = 10.42, logK3 = 9.44, logK4 = 8.28 olarak bulundu. Yine potansiyometrik olarak Irwing-Rossotti oluşum sabitleri ise alüminyum(III) için logK1 = 23.02, bakır(II) için logK1 = 19.92 olarak bulundu. Oluşan komplekslerin koşullu oluşum sabitleri hesaplandı ve buradan kompleksleşmenin ortaya çıktığı pH aralıkları bulundu. Alüminyum(III) ve bakır(II) komplekslerinin bileşimleri ligand/ metal = 1/ 1 olarak bulundu. Potansiyometrik sonuçlara dayanarak kuersetin/ metal = 1/ 1 oranıyla hazırlanan kompleksler için ters fazlı yüksek basınçlı sıvı kromatografisi ile tayin yöntemi gerçekleştirildi. Çalışmada hareketli faz olarak 0.01 M HCLO4 / 8.33 x 10-5 M Kuersetin çözeltisi (metanolde) (40/ 60), XTerra RP18, 5 μm, 4.6 x 150 mm özellikte kolon, λ = 373 ve 420 nm (bant genişliği 4nm) kullanıldı. Kuersetin-bakır(II) kompleksinin alıkonma zamanı TR = 3.706, kuyruklanma faktörü 0.957 dir. Kuersetin-alüminyum(III) kompleksinin alıkonma zamanı TR = 3.606, kuyruklanma faktörü 1.303 dür. Geliştirilen yöntem, yüksek miktarda kuersetin içerdiği bilinen Allium cepa (soğan) kabuklarıyla uygulanmıştır. Asit hidroliz sonucu kuersetin aglikon formu elde edilmiştir ve buradan kuersetinin alüminyum(III) ve bakır(II) kompleksleri katı halde çöktürülmüştür. Analiz edilen komplekslerin kromatogramları ve spektrumlarının standart olarak hazırlanan kursetin alüminyum(III) ve bakır(II) kompleksleri ile uyum içerisinde olduğu anlaşılmıştır. Anahtar sözcükler: Flavonoidler, flavonoidlerin metal kompleksleri, kromatografi, kuersetin, potansiyometri.
DETERMINATION of FLAVONOIDS and METAL COMPLEXES of FLAVONOIDS SIMULTANEOUSLY by HIGH PRESSURE LIQUID CHROMATOGRAPHY In this study, the stability constants of the aluminium(III) and copper(II) complexes of quercetin were determined potentiometrically by using Calvin-Bjerrum and Irwing Rossotti methods. The protonation constants of quercetin were found: logK1 = 11.15, logK2 = 10.42, logK3 = 9.44, logK4 = 8.28. For aluminium complex formation constant were found by Irwing-Rossotti method: logK1 = 23.02 and for copper complex: logK1 = 19.92. Conditional formation constants for complexes are calculated and pH ranges that complexation occurred are found. From the results, the components of quercetin / metal complexes are given as 1/ 1 both aluminium(III) and copper(II). A reversed phase high pressure liquid chromatographic method was developed for the metal complexes which were prepared according to potentiometric results with the ratio quercetin / metal = 1. Mobile phase was 0.01 M HCLO4 / 8.33 x 10-5 M quercetin in methanol (40/ 60), column was XTerra RP18, 5 μm, 4.6 x 150 mm, detector was Diode Array Detector at λ = 373 ve 420 nm (band width 4nm). For quercetin - aluminium(III) peak retention time was 3.706 and tailing factor was 0.957. For quercetin - copper(II) peak retention time was 3.606 and tailing factor was 1.303. The method was applied to Allium cepa (onion) dry outer skin that contains high amounts of quercetin. After acid hydrolsis, quercetin, aglycone was obtained, and quercetin complexes with aluminium(III) and copper(II) were precipitated. Solid complexes were analysed. It was found that, obtained chromatograms and spectrums are in accordance with the standart quercetin complexes with aluminium(III) and copper(II). Key words: Chromatography, flavonoids, metal complexes of flavanoids, potantiometry, quercetin.
DETERMINATION of FLAVONOIDS and METAL COMPLEXES of FLAVONOIDS SIMULTANEOUSLY by HIGH PRESSURE LIQUID CHROMATOGRAPHY In this study, the stability constants of the aluminium(III) and copper(II) complexes of quercetin were determined potentiometrically by using Calvin-Bjerrum and Irwing Rossotti methods. The protonation constants of quercetin were found: logK1 = 11.15, logK2 = 10.42, logK3 = 9.44, logK4 = 8.28. For aluminium complex formation constant were found by Irwing-Rossotti method: logK1 = 23.02 and for copper complex: logK1 = 19.92. Conditional formation constants for complexes are calculated and pH ranges that complexation occurred are found. From the results, the components of quercetin / metal complexes are given as 1/ 1 both aluminium(III) and copper(II). A reversed phase high pressure liquid chromatographic method was developed for the metal complexes which were prepared according to potentiometric results with the ratio quercetin / metal = 1. Mobile phase was 0.01 M HCLO4 / 8.33 x 10-5 M quercetin in methanol (40/ 60), column was XTerra RP18, 5 μm, 4.6 x 150 mm, detector was Diode Array Detector at λ = 373 ve 420 nm (band width 4nm). For quercetin - aluminium(III) peak retention time was 3.706 and tailing factor was 0.957. For quercetin - copper(II) peak retention time was 3.606 and tailing factor was 1.303. The method was applied to Allium cepa (onion) dry outer skin that contains high amounts of quercetin. After acid hydrolsis, quercetin, aglycone was obtained, and quercetin complexes with aluminium(III) and copper(II) were precipitated. Solid complexes were analysed. It was found that, obtained chromatograms and spectrums are in accordance with the standart quercetin complexes with aluminium(III) and copper(II). Key words: Chromatography, flavonoids, metal complexes of flavanoids, potantiometry, quercetin.