Mass Spectrometry-Based Metabolomic Analysis of Guava (Psidium guajava L.) and Its Antidiabetic Active Components

Abstract

In this study, metabolite profiling of guava was performed using mass spectrometry (MS)-based instruments, gas chromatography time-of-flight mass spectrometry (GC-TOF-MS) and liquid chromatography mass spectrometry (LC-MS), and bioactive compounds related to antidiabetic effects were examined combined with multivariate analysis. Terpene hydrocarbons and C6 compounds were the most abundant volatile metabolites in guava fruits and leaves. In particular, α-pinene and β-caryophyllene were dominant terpene hydrocarbons in both guava samples. Also, diverse C6 compounds such as 3-hexene-1-ol, (E)-2-hexen-1-ol, (Z)-3-hexenal, 2-hexenal, ethyl hexanoate and (Z)-3-hexenyl acetate were major metabolites. Non-targeted analysis of non-volatile metabolites in guava fruits was employed using two different data processing software, ChromaTOF and AMDIS. After data processing, metabolites of guava fruits could be separated by principal component analysis (PCA) according to not only different parts (flesh and peel) but also different ripening stages. Malic acid, aspartic acid, and glucose were the major metabolites distinguishing the different parts, while serine, fructose, and sucrose were related to the ripening of guava fruits. Metabolite profiling of guava leaves according to different harvest periods was also determined by metabolomic approach, and their antidiabetic activities were evaluated in vitro and in vivo studies. Some polyphenol compounds were major metabolites that discriminated guava leaves at different harvest times. In particular, guava leaves harvested in May (GVL1), the youngest leaves, possessed the highest level of polyphenols such as gallic acid, epicaechin, and quercetin. In addition, total phenolic content (TPC), α-glucosidase inhibition activity (AGIA) and oral glucose tolerant test (OGTT) were measured to investigate the antibiabetic effects of guava leaves. As a different mechanism of antidiabetic effects, the reduction effect of dicarbonyl compounds was also examined. In order to investigate the relationship between the metabolites and antidiabetic activities of guava leaves according to different harvest periods, partial least square regressions (PLSR) were carried out. Metabolites of GVL1 were significantly correlated with TPC and AGIA. In particular, polyphenol compounds such as epicatechin, quercetin, caffeic acid and quinic acid of GVL1 could be considered as bioactive compounds related to antidiabetic effects.;본 연구에서는 mass spectrometry에 근거한 기기인 GC-TOF-MS와 LC-MS를 이용하여 구아바 대사체들을 프로파일링 하고, 다변량통계분석을 적용하여 항당뇨 효과와 관련된 생리활성 성분을 고찰하였다. 구아바 과일과 잎의 휘발성 대사체 분석 결과, terpenes계 성분들과 C6 성분들이 다수 동정 되었으며, 특히, α-pinene과 β-caryopyllene은 과일과 잎의 주요 terpene계 성분으로 확인되었다. 또한, C6 화합물인 ethyl hexanoate, (Z)-3-hexenyl acetate, 3-hexene-1-ol, (E)-2-hexen-1-ol, (Z)-3-hexenal, 2-hexenal도 주요화합물로 동정되었다. Non-targeted 비휘발성 대사체 분석을 위하여 두 종류의 데이터 프로세싱 소프트웨어인 ChromaTOF와 AMDIS (Automated Mass Spectral Deconvolution and Identification System)를 사용하였으며, 그 결과, 주성분 분석 (principal component analysis, PCA)에 의해 구아바 과일의 다른 부위 (과피, 과육) 및 성숙 시기가 구분되었다. Malic acid, aspartic acid, glucose는 구아바 과육 및 과피를 구분 짓는 주요 대사체들로, serine, fructose, sucrose는 구아바의 성숙과 관련된 대사체들로 동정되었다. 또한, metabolomic 접근을 통한 시기별 구아바 잎의 대사체 프로파일링 및 생체 내∙외 항당뇨 효과 실험이 수행되었다. 시기별 구아바 잎을 구분 짓는 주요 대사체들로는 일부 폴리페놀 성분들이 확인되었으며, 특히, 5월에 수확된 구아바 잎은 quercetin, epicatechin, gallic acid와 같은 폴리페놀 성분들의 함량이 높게 나타났다. 한편, 구아바 잎의 항당뇨 효과를 알아보기 위하여 total phenolic content (TPC), α-glucosidase inhibition activity (AGIA), oral glucose tolerant test (OGTT)가 측정되었다. 또한, 항당뇨 효과의 다른 기전으로 dicarbonyl compounds의 저감화 효과가 확인되었다. 시기별 구아바 잎의 대사체와 항당뇨 효과와의 상관관계를 알아보기 위해서는 부분최소평방회귀분석 (Partial least square regression, PLSR)이 수행되었다. 첫번째 시기의 구아바 잎의 대사체와 total phenolic content (TPC), α-glucosidase inhibition activity (AGIA)는 유의적인 상관관계를 보였으며. 특히, 첫번째 시기의 구아바 잎의 epicatechin, quercetin, caffeic acid, quinic acid는 항당뇨 효과와 관련된 주요 바이오마커로 사료된다.