Publication: Türkiye’deki ceviz ağaçlarında bulunan ellajik asit ve gallik asidin hızlandırılmış solvent ekstraksiyonu ile izolasyonu ve miktar tayini
Abstract
Bu çalışmada Türkiye’deki ceviz ağaçlarından örnekler alınarak hızlandırılmış solvent ekstraksiyonu (ASE) uygulanmış, örneklerin içerdikleri ellajik asit (EA) ve gallik asit (GA) miktarları ters fazlı yüksek performanslı sıvı kromatografisi ile tayin edilmiştir. Ellajik Asit ve Gallik Asit polifenolik bileşiklerdir. İnsan sağlığı üzerinde birçok olumlu etkisi olan ellajik asidin en etkili özelliği kanser oluşumlarını engellemesidir. Özellikle akciğer, gırtlak, karaciğer, deri ve kolon kanserinde birçok araştırma yapılmıştır. Çalışmada hareketli faz bileşenleri olarak A: 0,025M KH2PO4 / H3PO4 tampon çözeltisi (pH: 3,0) ve B: Asetonitril (ACN) kullanılmıştır. Bu iki çözücü ile farklı dereceli elüsyon uygulamaları yapılmıştır. En uygun dereceli elüsyon sistemi EA pik alıkonma zamanı ve en yakın piklerin ayrılmaları göz önüne alınarak belirlenmiştir. Yeni bir ters fazlı yüksek performanslı sıvı kromatografisi (RP-HPLC) dereceli elüsyon yöntemi geliştirilmiştir. RP-HPLC sistemi: Phenomenex Luna C18 (2) 4,6 mm x 250 mm, 5 µm kolon, bant genişliği 4 nm, λ=255 nm dalga boyu ve diod dizi dedektör (DAD) kullanılmıştır. Beş farklı örnekle çalışılmıştır. 1. Ceviz ağacı gövdesi atığı (Rende), 2. Yaklaşık 70 yıllık ceviz ağacı kabuğu (K-70), 3. Yaklaşık 100 yıllık ceviz ağacı kabuğu (K-100), 4. Şebin türü ceviz ağacı filizi 5. Pedro türü ceviz ağacı filizi. Bu örneklerin ekstraksiyonunda Hızlandırılmış Solvent Ekstraksiyonu cihazı (ASE100) kullanılmıştır. Ektraksiyon koşulları çözücü metanol-su (90:10); sıcaklık 100˚C, statik zaman 1 dakika, temizleme hacmi % 50, yıkama zamanı 60 saniye, statik devir 3 olarak ayarlanmıştır. Kalibrasyon eğrisi y=189,904x + 1,101 r2: 0,9968. Geri kazanım % 35-43 olarak ceviz ağacı gövdesi atığı (rende) içinde hesaplanan EA miktarı % 21,65 bağıl standart sapma değeri ile 39,3 mg/ 100g olarak bulunmuştur. Tayin ve gözleme sınırları sırasıyla 13 ng.mL-1 (sinyal/ gürültü: 10/ 1) ve 60 ng mL-1 (sinyal/ gürültü: 3/ 1) olarak belirlendi. Ellajik Asit, Gallik Asit, hızlandırılmış çözücü ekstraksiyonu, miktar tayini, yüksek performanslı sıvı kromatografisi.
THE ISOLATION AND THE DETERMINATION OF ELLAGIC ACID AND GALLIC IN TURKEY’S WALNUT TREE BY ACCELERATED SOLVENT EXTRACTION In this study, the samples were collected from Turkey’s walnut trees, extracted by accelerated solvent extractor and determined the Ellagic acid (EA) and the Gallic Acid (GA) amounts by applying reverse phase-high performance liquide chromatography (RP-HPLC). Ellagid Acid ve Gallic Acid are poliphenolic compounds. Ellagic Acid plays significant role in human health by preventing the impovement of cancer. A number of studies have been reported on lung, throat, skin and colorectal cancer about the healing of EA. Accelerated Solvent Extractor (ASE100) was used for the extraction of the samples. Conditions for the extraction were set as: methanol-water 90-10; temperature 100˚C; static time 1 min; cleaning volume 50 %; cleaning time 60 sec; static cycle 3 cycles. The mobile phase components were A: 0,025M KH2PO4 / H3PO4 buffer solution (pH: 3,0) and B: Acetonitril (ACN). Various gardient elution systems were tried and the proper system was choosen considering the retention times and the resolution by using the A and B. The RP-HPLC system consisted of Phenomenex Luna C18 (2) 4,6 mm x 250 mm, 5 µm column and diode detector (DAD). The band width was 4 nm and the wavelength λ=255 nm. The five samples were studied: Walnut tree’s trunks; approximately 70 year-old walnuts tree’s bark (K-70); approximately 100 year-old walnut tree’s bark (K-100); buds of Şebin and Pedro walnuts. The constructed calibration curve was y=189,904x + 1,101 r2: 0,9968. The recovery was founded 35-43 %. The calculated amounts of EA the in the walnut tree’s trunks was founded 39,3 mg/ 100g with the RSD of % 21,65. The limit of quantification (LOQ) and limit of dedection (LOD) values were dedected respectively as 13 ng.mL-1 (signal/ noise: 10/ 1) and 60 ng mL-1 (signal/ noise: 3/ 1). Key words: Accelerated solvent extraction, determination, Ellagic Acid, Gallic Acid, high performance liquide chromatography.
THE ISOLATION AND THE DETERMINATION OF ELLAGIC ACID AND GALLIC IN TURKEY’S WALNUT TREE BY ACCELERATED SOLVENT EXTRACTION In this study, the samples were collected from Turkey’s walnut trees, extracted by accelerated solvent extractor and determined the Ellagic acid (EA) and the Gallic Acid (GA) amounts by applying reverse phase-high performance liquide chromatography (RP-HPLC). Ellagid Acid ve Gallic Acid are poliphenolic compounds. Ellagic Acid plays significant role in human health by preventing the impovement of cancer. A number of studies have been reported on lung, throat, skin and colorectal cancer about the healing of EA. Accelerated Solvent Extractor (ASE100) was used for the extraction of the samples. Conditions for the extraction were set as: methanol-water 90-10; temperature 100˚C; static time 1 min; cleaning volume 50 %; cleaning time 60 sec; static cycle 3 cycles. The mobile phase components were A: 0,025M KH2PO4 / H3PO4 buffer solution (pH: 3,0) and B: Acetonitril (ACN). Various gardient elution systems were tried and the proper system was choosen considering the retention times and the resolution by using the A and B. The RP-HPLC system consisted of Phenomenex Luna C18 (2) 4,6 mm x 250 mm, 5 µm column and diode detector (DAD). The band width was 4 nm and the wavelength λ=255 nm. The five samples were studied: Walnut tree’s trunks; approximately 70 year-old walnuts tree’s bark (K-70); approximately 100 year-old walnut tree’s bark (K-100); buds of Şebin and Pedro walnuts. The constructed calibration curve was y=189,904x + 1,101 r2: 0,9968. The recovery was founded 35-43 %. The calculated amounts of EA the in the walnut tree’s trunks was founded 39,3 mg/ 100g with the RSD of % 21,65. The limit of quantification (LOQ) and limit of dedection (LOD) values were dedected respectively as 13 ng.mL-1 (signal/ noise: 10/ 1) and 60 ng mL-1 (signal/ noise: 3/ 1). Key words: Accelerated solvent extraction, determination, Ellagic Acid, Gallic Acid, high performance liquide chromatography.