역상 고속 액체 크로마토그래피에 의한 식품 중 수용성 비타민 분석을 위한 전처리법의 비교

Comparision of Preparation Methods for Water Soluble Vitamin Analysis in Foods by Reversed-Phase High Performance Liquid Chromatography

  • 발행 : 2002.04.01

초록

식품에 함유된 수용성 비타민의 분석을 위하여 식품 중 단백질, 탄수화물 등과 결합하고있는 수용성 비타민을 추출 과정에서 파괴를 최소화하기 위하여 지금까지 통상적으로 행하여온 어떤 화학적 처리도 병용하지 않고 단백질 분해효소인 bromelain과 소의 췌장에서 추출한 protease 및 ${\alpha}-amylase$를 이용하여 결합을 절단하는 순수한 효소처리방법만을 이용하여 추출하였다. 추출된 비타민 시료용액을 $C_{18}$ Sep-Pak solid phase extraction column으로 정제하여 추출된 비타민을 한 column상에서 한번에 모두 역상 HPLC로 분석하는 방법을 개발하였다. 이 방법으로 표준 수용성 비타민의 회수율과 돼지고기 및 감자 중의 수용성 비타민분석 결과와 이 들 식품에 수용성 비타민을 강화하였을 때 강화 비타민의 회수율을 측정한 결과를 A.O.A.C.방법에 따라 추출 정제하여 분석한 결과와 비교하였다. Bromelain 처리와 A.O.A.C.법에서 표준수용성 비타민 중 folic acid가 가장 낮은 37.63%와 48.18%를 나타내었고 protease 처리에서는 pyridoxal이 가장 낮은 75.45%를 나타내었다. Niacinamide를 제외하고는 효소처리법이 A.O.A.C.법보다 높은 회수율을 보였다. 돼지고기 중 수용성비타민은 효소 처리방법에서 aminobenzoic acid, pyridoxal, pyridoxine, niacinamide 및 thiamin 등 6가지가 검출되었고 $C_{18}$ Sep-Pak column에 의한 정제에도 불구하고 불순 peak들이 많이 나왔으나 정량에 방해되는 불순 peak는 없었고 A.O.A.C.방법에 의한 추출 정제 시 niacinamide와 thiamin정량에 있어서 불순 peak에 의하여 정확한 정량이 불가능하였다. 돼지고기에서 특히 단백질과 공유결합을 형성하고 있는 비타민 $B_6$의 경우 문헌에 보고된 값들 보다 상당히 높은 값으로 정량되어 단백질 함량이 높은 시료에 대하여 별도의 정제과정이 추가로 필요한 것으로 나타났다. 비타민 강화 돼지고기 중의 강화 비타민의 회수 율은 표준 비타민 보다 회수율이 높고 정확하였다. 감자의 경우 protease처리에서 aminobezoic acid, pyridoxal, pyridoxine 및 thiamin 등 4가지가 검출되었고 bromelain 처리에서는 pyridoxal이 검출되지 않았다. 효소처리법에서 돼지고기와 마찬가지로 $C_{18}$ Sep-Pak column에 의한 정제로도 불순 peak가 많았으나 정량에 방해되는 불순 peak는 없었다. A.O.A.C.방법에서 pyridoxamine과 thiamin정량에서 불순 peak로 인하여 정확한 정량이 불가능하였다. 비타민 $B_6$의 경우 돼지고기와 달리 문헌에 나타난 값들과 비슷한 결과를 얻었는데 이는 단백질 함량이 낮은 관계로 판단된다. 비타민 강화 감자에서 강화 비타민의 회수 율도 돼지고기와 마찬가지로 상당히 정확하였고 A.O.A.C.법보다 높게나왔다.

Owing to a need for simple extraction and purification for analysis of water soluble vitamins in food samples by RP-HPLC with UV-detector, the methods of bromelain and protease hydrolysis and $C_{18}$ Sep-Pak solid phase extraction were employed. The recoveries of standard water soluble vitamins by the bromelain and protease hydrolysis and $C_{18}$ Sep-Pak solid phase extraction were significantly high compared to AOAC methods in most of vitamins. The contents of pyridoxal determined with protest in the pork was similar, but in the bromelain hydrolysis and AOAC method, was high compared to the results of reference. The niacinamide, thiamin and riboflavin determined with bromelain and protease hydrolysis showed similar values to the results of references. In the potato, pyridoxamine was detected in the AOAC method, which was not detected in the bromelain and protease hydrolysis methods. Pyridoxal contents in the protease hydrolysis and AOAC methods were very similar to the results of references. The recoveries of fortified standard vitamins in food samples were significantly high and accurate compared to those of AOAC methods. The extraction and purification with $C_{18}$ Sep-Pak solid extractor might be considered superior method for the determination of water soluble vitamins in food samples.

키워드

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