Evaluation of cellular antioxidant components of honeys using UPLC-MS/MS and HPLC-FLD based on the quantitative composition-activity relationship

doi:10.1016/j.foodchem.2019.04.105

Food Chemistry

Volume 293, 30 September 2019, Pages 169-177

https://doi.org/10.1016/j.foodchem.2019.04.105 Get rights and content

Highlights

•
The CAA of different honeys were evaluated comprehensively.
•
The CAA markers in honey were firstly explored based on the QCAR.
•
The antioxidant research on fennel honey was firstly investigated.
•
Phenolic acids, flavonoids and free amino acids in different honeys were analyzed.
•
Isosakuranetin, cryptochlorogenic acid and methionine were firstly found in honey.

Abstract

Antioxidant molecules in honey contributed to various biological effects, but antioxidant components markers in honey are required to be investigated further. Phenolic compounds, flavonoids and free amino acids were analyzed using UPLC-MS/MS and HPLC-FLD from 39 honey samples, in which fennel honey was firstly investigated. Based on the quantitative composition-activity relationship, the cellular antioxidant activity (CAA) assay of various honeys is closely related with the interaction of some phenolic compounds (isoferulic acid, 3,4-dihydroxy benzoic acid, 4-hydroxy benzoic acid, chlorogenic acid, caffeic acid, gallic acid, cryptochlorogenic acid, p-coumaric acid, salicylic acid), flavonoids (isosakuranetin, sakuranetin, pinocembrin, vitexin, taxifolin, galangin, luteolin, chrysin) and free amino acids (Tyr, Gly, Ile, Glu, Val, Phe, Leu, Asp, His, Pro, Ala). The results would be beneficial for the understanding of the nutritional values, exploitation and utilization of honeys with different floral origins, further contributable to the market development and consumption choice of honey.

Introduction

Honey is a natural substance well known for its various biological activities, e.g. antioxidant (Pichichero et al., 2009, Di Marco et al., 2018, Zhou et al., 2012), antimicrobial (Leyva-Jimenez et al., 2019, Fyfe et al., 2017, Pasias et al., 2018), and anti-inflammatory activities (Kassim et al., 2010, Abdelrahman et al., 2018, Zaidi et al., 2019). Carbohydrates comprise 95% of honey dry weight, especially mono- and di-saccharides such as glucose and fructose in larger quantities. There are also various proteins, minerals, amino acids, phenolic compounds and other minor components in honey, which are closely related to their botanical origin, processing and environmental conditions (Gašić et al., 2014, Chen et al., 2017).

Oxidative stress occurs when the level of oxidation exceeds the antioxidant defenses, which produces oxidative damage that exerts an influence on various physiological functions. Oxidation is caused by reactive oxygen species and free radicals in cellular systems, which are involved in senescence and incidence of cardiovascular disease, diabetes mellitus, and cancer (Rahal et al., 2014). There are various antioxidant molecules, containing phenolic acids, flavonoids and amino acids (Pérez, Iglesias, Pueyo, González, & de Lorenzo, 2007). In most cases, chemical-based methods are applied to evaluate the antioxidant activity of honey, e.g. Ferric reducing antioxidant power (FRAP) assay, 2,2-Diphenyl-1-picrylhydrazyl (DPPH) assay, 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assay and Oxygen radical absorbance capacity (ORAC) method (Bueno-Costa et al., 2016, Can et al., 2015, Gašić et al., 2014). However, the cellular antioxidant activity assay (CAA) not only quantifies bioactivities of antioxidant compounds in cultured cells, but also comprehensively considers the adsorption, distribution, metabolism, excretion and bioavailability of the antioxidant compounds. So, CAA method is more biologically representative than the chemical-based methods. Although CAA was reported for the antioxidant assessment of various fruits (Wolfe et al., 2008, Wolfe and Liu, 2007), it has not still been applied to the systematic research of honey except only one paper reported CAA for the phenolic extracts of buckwheat and manuka honey (Deng et al., 2018). Actually, more kinds of honey should be investigated for the systematic and comprehensive evaluation of the antioxidant activity of honey. Meanwhile, current available methods for antioxidant study of honey just simply determine the activity or analyze the chemical components. However, considering the antioxidant activity of honey is closely related with the interactions of multiple components, there are still challenges that need to holistically evaluate the cellular antioxidant activity of honeys with different floral and geographic origins, and comprehensively disclose the specific and effective antioxidant components in honey. Therefore, it is necessary to establish a quantitative composition-activity relationship (QCAR) to construct the integrative evaluation, and further discover contributing components in the antioxidant activity of honey (Zhang et al., 2018, Jiang et al., 2012).

In the present research, 39 honey samples from different floral origins (acacia, jujube, vitex, linden, fennel, buckwheat and manuka) were collected to analyze the bioactive compounds using ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) and high-performance liquid chromatography with fluorescence detection (HPLC-FLD) with pre-column derivatization. The antioxidant components and activity of fennel honey were firstly investigated. For the first time, CAA assay and QCAR were combined to systematically and comprehensively evaluate the antioxidant markers in honeys, which may facilitate the promotion of the nutritional values and the exploration and utilization of various honeys with different floral and geographic origins.

Section snippets

Honey samples

Acacia, jujube, vitex, linden, fennel, buckwheat, manuka honey samples were acquired from local beekeepers or purchased from manufacturers. All samples were stored at 4 °C. The collection sites were presented in Table 1.

Chemical reagents

Analytical standards 3,4-dihydroxy benzoic acid (98.2%), 4-hydroxy benzoic acid (99.7%), chlorogenic acid (96.0%), caffeic acid (99.2%), syringic acid (96.4%), p-coumaric acid (99.6%), sinapic acid (98%), ferulic acid (99.6%), quercetin (96.0%), quercetrin (98.3%), fisetin

Determination of TPC and TFC

TPC and TFC of 39 honey samples were determined, and the results indicated that TPC ranged from 9.15 (acacia honey) to 294 (buckwheat honey) mg GAE/100 g and TFC ranged from 6.85 (acacia honey) to 64.8 (buckwheat honey) mg QE/100 g, as shown in Table 1. Compared to the highest phenolic content (89.9 mg GAE/100 g) reported in manuka honeys among four floral honeys examined (Alzahrani et al., 2012), buckwheat honeys had the highest phenolic content (61.4–294 mg GAE/100 g), followed by fennel

Conclusion

In the present research, more antioxidant components of phenolic acids, flavonoids and free amino acids in honeys from different floral origins were analyzed using UPLC-MS/MS, HPLC-FLD, and isosakuranetin, cryptochlorogenic acid and methionine were firstly found in honeys. Phenolic acids, flavonoids and free amino acids in honeys varied, relating to the differential antioxidant competences and potential floral markers. CAA assay and QCAR were combined to systematically and comprehensively

Acknowledgements

The research was supported by the Grant from National Natural Science Foundation of China financially (No. 31501477).

Conflict of Interest.

The authors declare no conflict of interest.

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¹: Shi Shen and Jingbo Wang contributed equally to this work.

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Evaluation of cellular antioxidant components of honeys using UPLC-MS/MS and HPLC-FLD based on the quantitative composition-activity relationship

Highlights

Abstract

Introduction

Section snippets

Honey samples

Chemical reagents

Determination of TPC and TFC

Conclusion

Acknowledgements

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Biocatalysis and Agricultural Biotechnology

LWT - Food Science and Technology

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Talanta

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Analytica Chimica Acta

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European Journal of Integrative Medicine