Bioactive compounds, such as carotenoids, alkaloids, and phenolics, are well known because of their alleged health benefits when consumed regularly in a balanced healthy diet. Some well-documented ...bioactivities are antioxidant, antihypertensive, antihyperglycemic, antilipidemic, anti-obesity, anti-inflammatory, and antimicrobial capacities. Trying to associate the chemical composition of distinct sources and their bioactivity using in vitro methods, several assays have been developed, implemented, and optimised to recapitulate human physiological conditions. However, in most cases, pitfalls are apparent, and no single test tube-based assay can predict in vivo responses. The need for a more physiologically relevant cell-based method to evaluate the antioxidant capacity of putative antioxidants is apparent. Therefore, in this Review, the current state-of-the-art in food science and nutrition is aligned with cell biology/bioengineering approaches to propose combining in vitro digestion and absorption to obtain a bioavailable fraction containing antioxidants. Overall, human plasma, 2-dimensional human cell lines, such as erythrocytes, lymphocytes, hepatocytes, enterocytes and, ultimately, 3-dimensional spheroids (organoids) could be used as biologically relevant models to assess the antioxidant activity of compounds, foods, and nutraceuticals. This versatile approach is deemed suitable, accurate, reproducible, and physiologically relevant to evaluate the protective effects of antioxidants against ROS-mediated oxidation in vitro.
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•Chemical antioxidant activity does not reflect cellular physiological conditions.•Bioavailability and biotransformation of antioxidants are currently ignored.•In vitro digestion and absorption using Caco-2 co-culture should be used.•Cellular antioxidant activity should be performed using bioavailable fractions.•2D/3D human cell models must be used in antioxidant activity tests.
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•Tea catechins were studied for their combined antioxidant and cytotoxic effects.•EGCG presented the highest Cu2+ chelating ability.•67.4% ECG and 32.6% EGCG was the optimal ...combination (OPC)•OPC enhanced the anti-proliferative activity towards OVCAR-3, HEK293 and HFL1 cells.•OPC enhanced the bitterness and astringency of the chrysanthemum beverage model.
The effects of epigallocatechin gallate (EGCG), epigallocatechin (EGC) and epicatechin gallate (ECG) on the chemical and cell-based antioxidant activity, sensory properties, and cytotoxicity of a catechin-free model beverage were modeled using response surface methodology. Results showed that ECG presented the highest reducing capacity while EGCG presented the highest Cu2+ chelating ability. Binary interactions (EGCG/EGC and EGCG/ECG) had an additive effect on CUPRAC, DPPH and Cu2+ chelating ability. The mixture containing 67.4% ECG and 32.6% EGCG was the optimal combination of flavanols (OPC). In a beverage model – chrysanthemum tea – OPC enhanced the anti-proliferative activity in relation to OVCAR-3, HEK293 and HFL1 cells and decreased the intracellular generation of reactive oxygen species. OPC enhanced the bitterness and astringency of the beverage models impacting in a decrease in overall acceptance. The pasteurization process did not decrease the antioxidant activity and the flavanol concentration of the beverages.
Much research has been conducted to attest that different food and herbal extracts display functional properties in humans when consumed regularly as part of a balanced diet. However, most studies ...only relate one or some in vitro and/or in vivo functionalities of these extracts, without performing clinical trials to attest the alleged functionality. For instance, while some studies focus on the existence of statistical correlation between antioxidant activity and antidiabetic properties and the phenolic composition of a certain herb, others aim to assess the effects of different extraction methods (i.e., pulsed electric fields, ultrasound, and supercritical fluid extraction) on some selected in vitro functional properties.
Although these studies are essential for a better understanding of the extracts and foods consumed worldwide, they lack depth and, principally, practical application on consumer's health and well-being. In this scenario, in this article we propose an integration of multiple interlinked disciplines and between academia and food companies to elucidate the health-promoting properties of foods and extracts. In addition, we discuss and propose a multidimensional team to develop new functional foods.
Researchers should perform a wide variety of in vitro and in vivo tests to determine the toxicological effects, therapeutic dosage and to assess the physicochemical, chemical, and sensory properties of a developed food before stating its functionality. Complementarily, food scientists should never develop and attest in vivo functionality alone; rather, a highly active interconnection with related fields is required.
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•Functional foods have been studied and developed worldwide.•In vitro and in vivo studies are necessary to attest functionality of foods and extracts.•Functional food allegations must be based on scientific data.•The role of multidisciplinary studies is explained and integrated in this paper.
Tea is a typical processed beverage from the fresh leaves of Camellia sinensis Camellia sinensis (L.) O. Kuntze or Camellia assamica Camellia sinensis var. assamica (Mast.) Kitamura through different ...manufacturing techniques. The secondary metabolites of fresh tea leaves are mainly flavan‐3‐ols, phenolic acids, purine alkaloids, condensed tannins, hydrolysable tannins, saponins, flavonols, and their glycoside forms. During the processing, tea leaves go through several steps, such as withering, rolling, fermentation, postfermentation, and roasting (drying) to produce different types of tea. After processing, theaflavins, thearubigins, and flavan‐3‐ols derivatives emerge as the newly formed compounds with a corresponding decrease in concentrations of catechins. Each type of tea has its own critical process and presents unique chemical composition and flavor. The components among different teas also cause significant changes in their biological activities both in vitro and in vivo. In the present review, the progress of tea chemistry and the effects of individual unit operation on components were comprehensively described. The health benefits of tea were also reviewed based on the human epidemiological and clinical studies. Although there have been multiple studies about the tea chemistry and biological activities, most of existing results are related to tea polyphenols, especially (‐)‐epigallocatechin gallate. Other compounds, including the novel compounds, as well as isomers of amino acids and catechins, have not been explored in depth.
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•Fe2+ and Cu2+ chelating ability micro assays were standardized.•Coffees were analyzed for metal chelating ability, FRAP, DPPH and total phenolic content.•The methods presented a good ...repeatability and reproducibility (<7% RSD).•Cu2+ chelating ability was correlated to FRAP, DPPH and total phenolic content.
Aiming to standardize the experimental protocols to assess the ability to chelate Fe2+ and Cu2+ using 96-well microplates, we analyzed Brazilian coffees (n=20) as a study-case in relation to their antioxidant activity using conventional methods (DPPH and FRAP assays) and correlated the results with the total phenolic content (TPC) using bivariate and multivariate statistical approaches. Complementarily, we assessed the repeatability, reproducibility, recovery, and linearity of both methods. Data showed that the proposed assays presented a good repeatability and reproducibility (<7% RSD) and mean recovery values of 96.66% and 98.91% for the iron and copper assays, respectively. Both methods were linear in the range of 0–100mg EDTA equivalents/L. Cu2+-chelating ability was significantly correlated to FRAP, DPPH, and TPC, while sparse (p<0.05) correlations were obtained with Fe2+-chelating ability. Overall, both micro assays can be used to assess the ability of plant-based extracts to chelate Fe2+ and Cu2+in vitro.
The development of statistical software has enabled food scientists to perform a wide variety of mathematical/statistical analyses and solve problems. Therefore, not only sophisticated analytical ...methods but also the application of multivariate statistical methods have increased considerably. Herein, principal component analysis (PCA) and hierarchical cluster analysis (HCA) are the most widely used tools to explore similarities and hidden patterns among samples where relationship on data and grouping are until unclear. Usually, larger chemical data sets, bioactive compounds and functional properties are the target of these methodologies.
In this article, we criticize these methods when correlation analysis should be calculated and results analyzed.
The use of PCA and HCA in food chemistry studies has increased because the results are easy to interpret and discuss. However, their indiscriminate use to assess the association between bioactive compounds and in vitro functional properties is criticized as they provide a qualitative view of the data. When appropriate, one should bear in mind that the correlation between the content of chemical compounds and bioactivity could be duly discussed using correlation coefficients.
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•Chemometric tools are widely used for classification, calibration and exploratory issues.•Unsupervised statistical methods are used to study data structure and look for clusters of samples.•PCA and CA are the most widely used methods.•PCA and CA can be useful in studies regarding bioactive compounds in foods.•We criticize the indiscriminate use of PCA and CA.
Folin‐Ciocalteu colorimetric assay (FC) is the most widely used assay to estimate the total phenolic content in foods, beverages, herbs and other plant extracts, but many chemical compounds may act ...as interfering agents, producing inaccurate estimations of the real concentration of phenolic compounds in the matrix. Based on this limitation, the objective of this study was to compare, quantitatively, the Folin‐Ciocalteu and Prussian Blue (PB) assays in estimating the total phenolic content in purple grape juices (n = 20; Vitis labrusca L.) and teas (n = 25) from different botanical origins using 96‐well microplates. PB assay presented a low limit of detection (PB = 0.27 mg/L; FC = 0.25 mg/L) and quantification (PB = 0.92 mg/L; FC = 0.82 mg/L), showing its suitability in screening the total phenolic content in grape juices and teas. FC and PB assays presented a high association (P < 0.0001) for teas (r = 0.887) and grape juices (r = 0.923). The advantages of PB over FC assay are its simplicity, low time consumption (15 min reaction as compared to 60 min reaction for the FC assay), lower usage of reagents (solutions are prepared in a mM base), and higher selectivity. Additionally, PB assay was proven to be reproducible and repeatable and, therefore, may be used as an alternative to FC assay.
Practical application
Prussian Blue assay (PB) has been used as an alternative to Folin‐Ciocalteu assay (FC) to estimate the total content of phenolic compounds in herbs and some natural products. In our study we showed that the advantages of PB assay over FC are its simplicity, low time consumption (15 min reaction as compared to 60 min reaction for the FC assay), lower usage of reagents (solutions are prepared in a mM base) and higher selectivity as compared to FC assay. Additionally, PB assay was proven to be reproducible and repeatable and, therefore, may be used as an alternative to FC assay.
Statistical methods are important aids to detect trends, explore relationships and draw conclusions from experimental data. However, it is not uncommon to find that many researchers apply statistical ...tests without first checking whether they are appropriate for the intended application. The aim of this paper is to present some of the more important univariate and bivariate parametric and non-parametric statistical techniques and to highlight their uses based on practical examples in Food Science and Technology. The underlying requirements for use of particular statistical tests, together with their advantages and disadvantages in practical applications are also discussed, such as the need to check for normality and homogeneity of variances prior to the comparison of two or more sample sets in inference tests, correlation and regression analysis.
•The paper reviews the most used statistical methods in Food Science.•Normality, homoscedasticity, correlation, regression, censored data are explained.•Requirements for use of these tests, advantages and disadvantages are discussed.
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•Folin-Ciocalteu reagent and AlCl3 are used to assess the total phenolics and total flavonoids.•ABTS, CUPRAC, ORAC, DPPH and many other assays are used to assess the antioxidant ...activity.•Colorimetric methods present many pitfalls but can be used as screening tools.•LC–MS is recommended to quantify antioxidant compounds in food matrices.•In vivo tests and clinical studies are required to attest functionality of foods.
As many studies are exploring the association between ingestion of bioactive compounds and decreased risk of non-communicable diseases, the scientific community continues to show considerable interest in these compounds. In addition, as many non-nutrients with putative health benefits are reducing agents, hydrogen donors, singlet oxygen quenchers or metal chelators, measurement of antioxidant activity using in vitro assays has become very popular over recent decades. Measuring concentrations of total phenolics, flavonoids, and other compound (sub)classes using UV/Vis spectrophotometry offers a rapid chemical index, but chromatographic techniques are necessary to establish structure-activity. For bioactive purposes, in vivo models are required or, at the very least, methods that employ distinct mechanisms of action (i.e., single electron transfer, transition metal chelating ability, and hydrogen atom transfer). In this regard, better understanding and application of in vitro screening methods should help design of future research studies on ‘bioactive compounds’.
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