Comprehensive two-dimensional gas chromatography as a boosting technology in food-omic investigations.

This review focuses on the role that comprehensive two-dimensional gas chromatography can play within the investigation workflows of food-omics and related disciplines and subdisciplines, including food metabolomics, nutrimetabolomics, sensomics, and food safety. After a short introductory survey, discussing the intriguing context of system biology and integrationist approaches of investigation, the concepts of analytical dimensions and the key characteristics of comprehensive two-dimensional gas chromatography are introduced. Through a selection of relevant examples, the boosting role of comprehensive two-dimensional gas chromatography within food-omics is described, providing to the reader evidence of how comprehensive multidimensional separations based platforms have introduced new concepts and tools in the analytical measurement of complex biological samples.

Untargeted metabolomics with multivariate analysis to discriminate hazelnut (Corylus avellana L.) cultivars and their geographical origin.

BACKGROUND: In the present study a metabolomics-based approach was used to discriminate among different hazelnut cultivars and to trace their geographical origins. Ultra-high-pressure liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry (UHPLC-ESI/QTOF-MS) was used to profile phenolic and sterolic compounds. RESULTS: Compounds were identified against an in-house database using accurate monoisotopic mass and isotopic patterns. The screening approach was designed to discern 15 hazelnut cultivars and to discriminate among the geographical origins of six cultivars from the four main growing regions (Chile, Georgia, Italy, and Turkey). This approach allowed more than 1000 polyphenols and sterols to be annotated. The metabolomics data were elaborated with both unsupervised (hierarchical clustering) and supervised (orthogonal projections to latent structures discriminant analysis, OPLS-DA) statistics. These multivariate statistical tools allowed hazelnut samples to be discriminated, considering both 'cultivar type' and 'geographical origin'. Flavonoids (anthocyanins, flavanols and flavonols - VIP scores 1.34-1.49), phenolic acids (mainly hydroxycinnamics - VIP scores 1.35-1.55) together with cholesterol, ergosterol, and stigmasterol derivatives (VIP scores 1.34-1.49) were the best markers to discriminate samples according to geographical origin. CONCLUSIONS: This work illustrates the potential of untargeted profiling of phenolics and sterols based on UHPLC-ESI/QTOF mass spectrometry to discriminate hazelnut and support authenticity and origin. © 2019 Society of Chemical Industry.

Untargeted screening of the bound / free phenolic composition in tomato cultivars for industrial transformation.

BACKGROUND: Tomato is one of the most important agricultural crops and it is characterized by a wide bioactive compound profile. However, little information is reported on its comprehensive polyphenol profile. In this work, 13 commercial tomato cultivars for industrial transformation were screened by ultra-high-pressure liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry (UHPLC-QTOF-MS) for both free and bound phenolic profiles. Thereafter, the in vitro antioxidant activity of each cultivar was assessed by ferric reducing antioxidant power (FRAP) and oxygen radical absorbance activity (ORAC) assays. Multivariate statistics, i.e. orthogonal projection to latent structures discriminant analysis (OPLS-DA), were then used to model samples according to their distinct phenolic signatures, thus providing compounds that better discriminated between the distributions of the cultivars that were considered. RESULTS: More than 350 phenolic compounds could be identified across the samples that were considered: flavonoids (such as flavones and flavanols), hydroxycinnamic acids, lignans, and lower-molecular-weight phenolics were the most frequently observed classes of phenolics in tomato berries. Anthocyanins were the most abundant class among bound phenolics (being highest in the Leader F1 and Defender F1 cultivars), followed by tyrosols (mainly in Heinz cultivars). However, flavones and hydroxybenzoic acids were the most represented discriminant phenolics in the bound fraction. CONCLUSIONS: Untargeted metabolomics allowed significant differences in phenolic composition to be outlined across the tomato cultivars that were analyzed. Such differences were particularly evident regarding the free-to-bound phenolic ratio, hence allowing differences in the bioaccessibility of phenolics to be postulated. © 2019 Society of Chemical Industry.

Potential of LC Coupled to Fluorescence Detection in Food Metabolomics: Determination of Phenolic Compounds in Virgin Olive Oil.

A powerful chromatographic method coupled to a fluorescence detector was developed to determine the phenolic compounds present in virgin olive oil (VOO), with the aim to propose an appropriate alternative to liquid chromatography-mass spectrometry. An excitation wavelength of 285 nm was selected and four different emission wavelengths (316, 328, 350 and 450 nm) were simultaneously recorded, working therefore on "multi-emission" detection mode. With the use of commercially available standards and other standards obtained by semipreparative high performance liquid chromatography, it was possible to identify simple phenols, lignans, several complex phenols, and other phenolic compounds present in the matrix under study. A total of 26 phenolic compounds belonging to different chemical families were identified (23 of them were susceptible of being quantified). The proposed methodology provided detection and quantification limits within the ranges of 0.004-7.143 µg·mL(-1) and 0.013-23.810 µg·mL(-1), respectively. As far as the repeatability is concerned, the relative standard deviation values were below 0.43% for retention time, and 9.05% for peak area. The developed methodology was applied for the determination of phenolic compounds in ten VOOs, both monovarietals and blends. Secoiridoids were the most abundant fraction in all the samples, followed by simple phenolic alcohols, lignans, flavonoids, and phenolic acids (being the abundance order of the latter chemical classes logically depending on the variety and origin of the VOOs).

Multidisciplinary approach combining food metabolomics and epidemiology identifies meglutol as an important bioactive metabolite in tempe, an Indonesian fermented food.

This study introduces a multidisciplinary approach to investigate bioactive food metabolites often overlooked due to their low concentrations. We integrated an in-house food metabolite library (n = 494), a human metabolite library (n = 891) from epidemiological studies, and metabolite pharmacological databases to screen for food metabolites with potential bioactivity. We identified six potential metabolites, including meglutol (3-hydroxy-3-methylglutarate), an understudied low-density lipoprotein (LDL)-lowering compound. We further focused on meglutol as a case study to showcase the range of characterizations achievable with this approach. Green pea tempe was identified to contain the highest meglutol concentration (21.8 ± 4.6 mg/100 g). Furthermore, we identified a significant cross-sectional association between plasma meglutol (per 1-standard deviation) and lower LDL cholesterol in two Hispanic adult cohorts (n = 1,628) (ß [standard error]: -5.5 (1.6) mg/dl, P = 0.0005). These findings highlight how multidisciplinary metabolomics can serve as a systematic tool for discovering and enhancing bioactive metabolites in food, such as meglutol, with potential applications in personalized dietary approaches for disease prevention.

Exploring stingless bee honey from selected regions of Peninsular Malaysia through gas chromatography-mass spectrometry-based untargeted metabolomics.

Volatile organic compounds in honey are known for their considerable impact on the organoleptic properties of honey, such as aroma, flavor, taste, and texture. The type and composition of volatile organic compounds are influenced by entomological, geographical, and botanical origins; thus, these compounds have the potential to be chemical markers. Sixty-two volatile compounds were identified using gas chromatography-mass spectrometry from 30 Heterotrigona itama (H. itama) honey samples from 3 different geographical origins. Hydrocarbons and benzene derivatives were the dominant classes of volatile organic compounds in the samples. Both clustering and discriminant analyses demonstrated a clear separation between samples from distant origins (Kedah and Perak), and the volcano plot supported it. The reliability and predictability of the partial least squares-discriminant analysis model from the discriminant analysis were validated using cross-validation (R2 : 0.93; Q2 : 0.83; accuracy: 0.97) and the permutation test (p < 0.001), and the output depicted that the model is legitimate. In combination with the variable importance of projection (VIP > 1.0) and the Kruskal-Wallis test (p < 0.01), 19 volatile organic compounds (encompassed aldehydes, benzene derivatives, esters, hydrocarbons, and terpenoids) were sorted and named potent chemical markers in classifying honey samples from three geographical origins. In brief, this study illustrated that volatile organic compounds of stingless honey originated from the same bee species, but different geographical origins could be applied as chemical markers.

Superior high-efficiency and high-throughput volatile flavor extraction of Japanese fermented seasonings by solvent-assisted stir bar solid extraction with reverse extraction.

Fermented seasonings have pleasant flavors that stimulate our appetite. Their flavoring properties change depending on factors such as their materials and fermented conditions. Therefore, a comparative analysis of their flavor is important when evaluating their quality. However, seasonings contain high levels of various matrices such as sugars, proteins, lipids, and ethanol, making it difficult to extract aroma compounds efficiently from them. In this study, we verified a high-efficient and high-throughput volatile flavor analysis of fermented seasonings by solvent-assisted stir bar solid extraction (SA-SBSE) with reverse extraction. We applied SA-SBSE to Japanese fermented seasonings, soy sauce, miso (fermented beans), and mirin (sweet rice wine) and compared their profiles with those from other common extraction methods, headspace gas-solid-phase microextraction (HS-SPME), liquid extraction with solvent-assisted flavor evaporation (LE-SAFE), and conventional SBSE (C-SBSE). The aroma properties and profiles of extracts from SA-SBSE were close to those of the original sample, being similar to that of LE-SAFE. In addition, potent aroma compounds in each sample were extracted by SA-SBSE and LE-SAFE, which were far superior to those by C-SBSE. For quantification, SA-SBSE extracts showed a good standard curve by the standard addition method. We could quantify maltol, one of the most common potent aroma compounds in all samples, for various commercial samples by such high-throughput analysis.

Untargeted metabolomics and conventional quality characterization of rowanberry pomace ingredients in meatballs.

In this study, a rowanberry pomace defatted with supercritical CO2 (2%-AC), its ethanolic extract (1%-E) and extraction residue (2%-R), were tested in meatball preparation. The meatballs with 1%-E demonstrated the highest in vitro radical scavenging capacity. In the case of 1%-E the pH of meatballs was significantly lower compared to the control sample (P = 0.0132) on the 5-day. The lowest cooking loss was achieved when the meatballs contained mainly fibre-rich 2%-R. The UHPLC method detected 184 metabolites, including strong antioxidants, such as chlorogenic acids, 3',4'-methylenedioxy-5,7-dimethylepicatechin, hyperin, isoquercitrin. The 1%-E was particularly effective against the development of unpleasant off-flavours caused by carbonyl compounds. Consistently, the decrease in lipid oxidation, indicated by reduced 7-dodecenal and 2,4-heptadienal contents, has been observed following the addition of rowanberry extract to meatballs. Metabolomics coupled with conventional quality evaluations provided a deeper understanding of the potential utilization and valorisation of different rowanberry pomace extracts as meat ingredients.

Improvement of the functional value of green soybean (edamame) using germination and tempe fermentation: A comparative metabolomics study.

Green soybean, also known as edamame, is a legume with high nutritional and functional value. Despite its growing popularity and potential health benefits, the functionality of green soybean has not been thoroughly studied. Previous research on the functionality of green soybean has largely focused on a limited number of specific, well-studied, bioactive metabolites, without comprehensively investigating the metabolome of this legume. Additionally, very few studies have explored the improvement of the functional value of green soybean. This study aimed to investigate the metabolome profile of green soybean, identify bioactive metabolites, and to further explore the potential improvement of the identified bioactive metabolites using germination and tempe fermentation. A total of 80 metabolites were annotated from green soybean using GC-MS and HPLC-PDA-MS. Among them, 16 important bioactive metabolites were identified: soy isoflavones daidzin, glycitin, genistin, malonyl daidzin, malonyl genistin, malonyl glycitin, acetyl daidzin, acetyl genistin, acetyl glycitin, daidzein, glycitein, and genistein, as well as other metabolites including 3,4-dihydroxybenzoic acid, 3-hydroxyanthranillic acid, 3-hydroxy-3-methylglutaric acid (meglutol), and 4-aminobutyric acid (GABA). Germination and tempe fermentation techniques were employed to potentially improve the concentrations of these bioactive metabolites. While showing improvements in amino acid contents, germination process did not improve bioactive metabolites significantly. In contrast, tempe fermentation was found to significantly increase the concentrations of daidzein, genistein, glycitein, acetyl genistin, acetyl daidzin, 3-hydroxyanthranillic acid, and meglutol (>2-fold increase with p < 0.05) while also improving amino acid levels. This study highlights the potentials of germination and fermentation to improve the functionality of legumes, particularly green soybean.

Wakame replacement alters the metabolic profile of wheat noodles after in vitro digestion.

The development of nutritional noodles of high quality has become a new hotspot of research in the area of food science. Since wakame is edible seaweed rich in dietary fiber and proteins and rarely found in ordinary noodle, this study investigated the release of metabolites, the texture quality, and the rheological properties of wakame noodle, as well as the mechanism by which extruded wakame flours can influence noodle texture and viscoelasticity through digestion. Basically, nuclear magnetic resonance spectra were applied to identify the 46 metabolites including amino acids, saccharides, fatty acids, and other metabolites. Both PCA and OPLS-DA model showed fit goodness and good predictivity, which were assessed the increasing release of most metabolites. Structural studies discussed the effects on the enhancement of interlinkage with gluten matrix and protein matrix, which were validated via the decreasing instantaneous compliance J0 (1.64 × 10-5 to 0.16 × 10-5 Pa-1). Wakame addition best matched the physiochemical properties of noodle, in terms of chewiness (99.10 vs 122.66 g.mm), gumminess (281.98 vs. 323.44 g), and gel strength (132.65 vs 173.95 kPa•s-1). Beyond the functional characteristics it contributes benefits like reduction of diet-related diabetes. As a consequence, the creation of personalized nutritious, healthy noodles will be an innovative route from a scientific viewpoint and an application standpoint.

Metabolomics as a tool for geographic origin assessment of roasted and green coffee beans.

Coffee is widely consumed across the globe. The most sought out varieties are Arabica and Robusta which differ significantly in their aroma and taste. Furthermore, varieties cultivated in different regions are perceived to have distinct characteristics encouraging some producers to adopt the denomination of origin label. These differences arise from variations on metabolite content related to edaphoclimatic conditions and post-harvest management among other factors. Although sensory analysis is still standard for coffee brews, instrumental analysis of the roasted and green beans to assess the quality of the final product has been encouraged. Metabolomic profiling has risen as a promising approach not only for quality purposes but also for geographic origin assignment. Many techniques can be applied for sample analysis: chromatography, mass spectrometry, and NMR have been explored. The data collected is further sorted by multivariate analysis to identify similar characteristics among the samples, reduce dimensionality and/or even propose a model for predictive purposes. This review focuses on the evolution of metabolomic profiling for the geographic origin assessment of roasted and green coffee beans in the last 21 years, the techniques that are usually applied for sample analysis and also the most common approaches for the multivariate analysis of the collected data. The prospect of applying a wide range of analytical techniques is becoming an unbiased approach to determine the origin of different roasted and green coffee beans samples with great correlation. Predictive models worked accurately for the geographic assignment of unknown samples once the variety was known.

The evolving role of the Caenorhabditis elegans model as a tool to advance studies in nutrition and health.

This review highlights the scope and relevance for using the nematode Caenorhabditis elegans for advancing human nutrition and health studies. This is in the context of changing global priorities by offering a simple tool to investigate the nutritional problems facing the world today. Existing research presented here shows the advantages of using C elegans to evaluate human eating behavior, obesity, and the molecular basis of nutrition and gustation. C elegans has been used to understand diet interventions and nutraceuticals and explain how these metabolites affect human health. The nematode has been used to investigate various aspects of the gut microbiome and selecting appropriate microbial strains for probiotic studies. Nematode research has interpreted mechanisms of host-pathogen interactions to control foodborne illness and examine diet-related aging and longevity. Future research using C elegans shows promise for evaluating space nutrition and immunity testing.

Xanthan gum modified fish gelatin and binary culture modulates the metabolism of probiotics in fermented milk mainly via amino acid metabolism pathways.

1H NMR combined with multivariate data analysis were applied to investigate the effects of fish gelatin (FG) addition and co-culture of Lactobacillus acidophilus LA-5 (La-5) and Bifidobacterium lactis BB-12 (Bb-12) on the growth and metabolic pathways of the probiotics themselves. The results showed that the addition of FG had no significant effects on the growth of probiotics, but co-culture did promote the growth of probiotics, especially for Bb-12 (up to 2 log CFU/mL). FG addition inhibited amino acids synthesis and TCA cycling in Lacticaseibacillus paracasei subsp. paracasei CASEI 431 (L431) to some extent. However, for the single La-5 strain, these pathways were promoted. As for mixed bacterial cultures, Bb-12 promoted amino acids metabolism, sugar transport and energy metabolism in La-5. These findings suggested that the metabolic profile of probiotic bacteria can be adequately explained by metabolic pathway analysis, which also provides theoretical guidance for the industrialization of functional fermented milk.

Metabolomic insight into the profile, in vitro bioaccessibility and bioactive properties of polyphenols and glucosinolates from four Brassicaceae microgreens.

In this study, four Brassicaceae microgreens species, namely kale, red cabbage, kohlrabi, and radish, were evaluated for their phytochemical compositions using spectrophotometric assays and untargeted metabolomics before and after in vitro gastrointestinal digestion. According to the in vitro spectrophotometric results, significant amounts of phenolics could be detected in each studied species, thus supporting the total antioxidant capacities recorded. Overall, metabolomics allowed annotating a total of 470 phytochemicals across the four Brassicaceae microgreens, either fresh or digested. Among polyphenols, flavonoids were the most represented class (180 compounds, including anthocyanins, flavones, flavonols, and other flavonoids), followed by phenolic acids (68 compounds, mainly hydroxycinnamic and hydroxybenzoic acids), non-flavonoid or phenolic acid-based structures (i.e., alkyl- and alkylmethoxy-phenols and tyrosol derivatives), and lignans. Also, 22 glucosinolates were annotated, including gluconapin glucoraphanin, glucobrassicin, and 4-hydroxyglucobrassicin. Noteworthy, significant differences could be observed in terms of bioaccessibility as a function of the phenolic class and the species considered. Overall, lignans exhibited the highest bioaccessibility values (14%), followed by tyrosol derivatives and flavonoids (on average, 9% and 8%, respectively). However, differences could be evidenced as a function of the species, with red cabbage having comparatively lower bioaccessibility values irrespective of the chemical class of bioactive considered. Similarly, bioaccessibility of glucosinolates significantly differed across species, ranging from 2% in kale to 43% in kohlrabi microgreens.

Polyphenol Exposure, Metabolism, and Analysis: A Global Exposomics Perspective.

Polyphenols are generally known for their health benefits and estimating actual exposure levels in health-related studies can be improved by human biomonitoring. Here, the application of newly available exposomic and metabolomic technology, notably high-resolution mass spectrometry, in the context of polyphenols and their biotransformation products, is reviewed. Comprehensive workflows for investigating these important bioactives in biological fluids or microbiome-related experiments are scarce. Consequently, this new era of nontargeted analysis and omic-scale exposure assessment offers a unique chance for better assessing exposure to, as well as metabolism of, polyphenols. In clinical and nutritional trials, polyphenols can be investigated simultaneously with the plethora of other chemicals to which we are exposed, i.e., the exposome, which may interact abundantly and modulate bioactivity. This research direction aims at ultimately eluting into atrue systems biology/toxicology evaluation of health effects associated with polyphenol exposure, especially during early life, to unravel their potential for preventing chronic diseases.

Comparative phytochemical profile of the elephant garlic (Allium ampeloprasum var. holmense) and the common garlic (Allium sativum) from the Val di Chiana area (Tuscany, Italy) before and after in vitro gastrointestinal digestion.

This study is aimed to comparatively investigate the phytochemical profiles, focusing on the nutritional and phytochemical properties of common garlic (Allium sativum L.; CG) and elephant garlic (EG) (Allium ampeloprasum var. holmense) collected from the Val di Chiana area (Tuscany, Italy). The results showed a lower amount of fibers, demonstrating a higher digestibility of the bulb, and sulfur-containing compounds in EG rather than in CG. Untargeted metabolomic profiling followed by supervised and unsupervised statistics allowed understanding the differences in phytochemical composition among the two bulbs, both as raw bulbs, processed following the in vitro gastrointestinal digestion process. Typical sulfur-containing compounds, such as alliin and N-gamma-glutamyl-S-allyl cysteine, could notably be detected in lower amounts in EG. EG maintains a distinct phytochemical signature during in vitro gastrointestinal digestion. Our findings support the distinct sensorial attributes of the bulbs.

Pigmented sorghum polyphenols as potential inhibitors of starch digestibility: An in vitro study combining starch digestion and untargeted metabolomics.

Polyphenols from five pigmented sorghum (PS) flours were in vitro evaluated as possible modulators of starch digestibility. White sorghum (WS) flour was used as control. Untargeted metabolomics depicted the phenolic composition of raw and cooked flours (obtained through heating at 100 °C for 30 min in water) highlighting differences in flavonoids and phenolic acids. Raw PS flours were characterized by greater tannin and kafirin contents when compared to WS, and, after cooking, PS flours had greater resistant starch (from 4.2 to 21.4 g /100 g dry matter), and lower starch hydrolysis index (HI) with respect to cooked WS. Multivariate statistics showed that flavonoids characterizing PS were the most discriminant compounds during the in vitro digestion. In addition, kafirin and total tannins content (on raw ingredients) along with the anthocyanin profiles (on cooked samples) were negative correlated with HI. Therefore, PS flours might be good candidates for the formulation of functional foods.

Impact of a Pitanga Leaf Extract to Prevent Lipid Oxidation Processes during Shelf Life of Packaged Pork Burgers: An Untargeted Metabolomic Approach.

In this work, the comprehensive metabolomic changes in pork burgers treated with different antioxidants, namely, (a) a control without antioxidants, (b) 200 mg/kg butylated hydroxytoluene (BHT), and (c) 250 mg/kg pitanga leaf extract (PLE, from Eugenia uniflora L.), each one packaged under modified atmosphere (80% O2 and 20% CO2) for 18 days storage at 2 ± 1 °C, were deeply studied. In particular, untargeted metabolomics was used to evaluate the impact of the antioxidant extracts on meat quality. The PLE phytochemical profile revealed a wide variety of antioxidant compounds, such as polyphenols, alkaloids, and terpenoids. Multivariate statistics (both unsupervised and supervised) allowed to observe marked differences in BHT and PLE burgers metabolomic profiles during storage. Most of the differences could be attributed to hexanoylcarnitine, 4-hydroxy-2-nonenal, 6-hydroxypentadecanedioic acid, 9S,11S,15S,20-tetrahydroxy-5Z,13E-prostadienoic acid (20-hydroxy-PGF2a), sativic acid, followed by glycerophospholipids. In addition, significant correlations (p < 0.01) were observed between thiobarbituric acid reactive substances and metabolites related to lipid oxidation processes. Therefore, the approach used showed a clear modulation of lipid oxidation, likely promoted by the plant leaf extract, thus confirming the ability of PLE to delay lipid oxidative phenomena during storage.

Metabolomics-Based Study of the Effect of Raw Materials to the End Product of Tempe-An Indonesian Fermented Soybean.

Tempe is a fermented soybean food from Indonesia, made by inoculating Rhizopus spp. onto cooked and dehulled soybean. Tempe has been a part of Indonesian culture since the 16th century and is now produced globally as a highly nutritious plant-based food. Despite a longstanding history on the production of tempe, very few studies have been reported to understand the effect of raw material to the end product metabolite composition. In this research, we applied GC/MS-based metabolite profiling to investigate the effect of various possible factors that might affect the final product (environmental factors, raw materials, and starter cultures). Representative samples were tempe produced by Indonesian industries, Japanese industries as well as laboratory made tempe. The results showed that both environmental factors and raw materials (soybean, water, and starter culture) contributed to the tempe metabolite profile. Here we found the possibility that starter cultures might play a greater role to determine the metabolite profiles compared to other tested factors. This research might provide useful insights for the larger scale industries to maintain the quality of tempe for the benefit of the consumers.

Phenolic profiling and in vitro bioactivity of Moringa oleifera leaves as affected by different extraction solvents.

In this work the (poly)-phenolic profile of Moringa oleifera leaves was comprehensively investigated through untargeted metabolomics, following a homogenizer-assisted extraction (HAE) using three solvent systems, i.e. methanol (HAE-1), methanol-water 50:50 v/v (HAE-2) and ethyl acetate (HAE-3). This approach allowed to putatively annotate 291 compounds, recording mainly flavonoids and phenolic acids. Thereafter, antioxidant capacity, antimicrobial activity and enzyme inhibition were assayed in the different extracts. HAE-1 extract showed the highest total phenolic content (31.84 mg/g), followed by HAE-2 (26.95 mg/g) and HAE-3 (14.71 mg/g). In addition, HAE-1 and HAE-2 extracts exhibited an expressive activity against Bacillus cereus and Listeria innocua. The HAE-2 leaf extract was characterized by the highest DPPH and ABTS values (being 49.55 and 45.26 mgTE/g), while ferric reducing antioxidant power was found to be higher in HAE-1 (58.26 mgTE/g). Finally, the enzyme inhibitory effects of M. oleifera leaf extracts were investigated against five enzymes, namely acetylcholinesterase (AChE), butyrylcholinesterase (BChE), tyrosinase, α-amylase and α-glucosidase. All of the tested extracts exhibited inhibitory effects on AChE and BChE with a higher activity for HAE-3 and HAE-1, whilst HAE-1 showed the higher impact on tyrosinase, glucosidase and amylase activities. Taken together, these findings suggest that M. oleifera leaf extracts are a good source of bioactive polyphenols with a potential use in food and pharma industries.