A serine carboxypeptidase-like acyltransferase catalyzes consecutive four-step reactions of hydrolyzable tannin biosynthesis in Camellia oleifera.

Hydrolyzable tannins (HTs), a class of polyphenolic compounds found in dicotyledonous plants, are widely used in food and pharmaceutical industries because of their beneficial effects on human health. Although the biosynthesis of simple HTs has been verified at the enzymatic level, relevant genes have not yet been identified. Here, based on the parent ion-fragment ion pairs in the feature fragment data obtained using UPLC-Q-TOF-/MS/MS, galloyl phenolic compounds in the leaves of Camellia sinensis and C. oleifera were analyzed qualitatively and quantitatively. Correlation analysis between the transcript abundance of serine carboxypeptidase-like acyltransferases (SCPL-ATs) and the peak area of galloyl products in Camellia species showed that SCPL3 expression was highly correlated with HT biosynthesis. Enzymatic verification of the recombinant protein showed that CoSCPL3 from C. oleifera catalyzed the four consecutive steps involved in the conversion of digalloylglucose to pentagalloylglucose. We also identified the residues affecting the enzymatic activity of CoSCPL3 and determined that SCPL-AT catalyzes the synthesis of galloyl glycosides. The findings of this study provide a target gene for germplasm innovation of important cash crops that are rich in HTs, such as C. oleifera, strawberry, and walnut.

New insights into the function of plant tannase with promiscuous acyltransferase activity.

Plant tannases (TAs) or tannin acyl hydrolases, a class of recently reported carboxylesterases in tannin-rich plants, are involved in the degalloylation of two important groups of secondary metabolites: flavan-3-ol gallates and hydrolyzable tannins. In this paper, we have made new progress in studying the function of tea (Camellia sinensis) (Cs) TA-it is a hydrolase with promiscuous acyltransferase activity in vitro and in vivo and promotes the synthesis of simple galloyl glucoses and flavan-3-ol gallates in plants. We studied the functions of CsTA through enzyme analysis, protein mass spectrometry, and metabolic analysis of genetically modified plants. Firstly, CsTA was found to be not only a hydrolase but also an acyltransferase. In the two-step catalytic reaction where CsTA hydrolyzes the galloylated compounds epigallocatechin-3-gallate or 1,2,3,4,6-penta-O-galloyl-ß-d-glucose into their degalloylated forms, a long-lived covalently bound Ser159-linked galloyl-enzyme intermediate is also formed. Under nucleophilic attack, the galloyl group on the intermediate is transferred to the nucleophilic acyl acceptor (such as water, methanol, flavan-3-ols, and simple galloyl glucoses). Then, metabolic analysis suggested that transient overexpression of TAs in young strawberry (Fragaria × ananassa) fruits, young leaves of tea plants, and young leaves of Chinese bayberry (Myrica rubra) actually increased the total contents of simple galloyl glucoses and flavan-3-ol gallates. Overall, these findings provide new insights into the promiscuous acyltransferase activity of plant TA.

Phytochemical profile and determination of cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity of aqueous and ethanolic extracts of Pseudobombax marginatum (A. St.-Hil.) A. Robyns.

Pseudobombax marginatum, popularly known as "embiratanha," is widely used by traditional communities as anti-inflammatory and analgesic agent. This study aimed to determine the phytochemical profile as well as cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity attributed to exposure to aqueous (AqEx) and ethanolic (EtEx) extracts of embiratanha bark. Phytochemical screening was conducted using thin-layer chromatography (TLC). Cell viability was analyzed using MTT assay with human mammary gland adenocarcinoma (MDA-MB-231) and macrophage (J774A.1) cell lines, exposed to concentrations of 12.5, 25, 50, or 100 µg/ml of either extract. For acute oral toxicity, comet assay and micronucleus (MN) tests, a single dose of 2,000 mg/kg of either extract was administered orally to Wistar rats. TLC analysis identified classes of metabolites in the extracts, including cinnamic acid derivatives, flavonoids, hydrolyzable tannins, condensed tannins, coumarins, and terpenes/steroids. In the cytotoxicity assay, the varying concentrations of extracts derived from embiratanha induced no significant alterations in the viability of MDA-MB-231 cells. The lowest concentration of EtEx significantly increased macrophage J774A.1 viability. However, the higher concentrations of AqEx markedly lowered macrophage J774A.1 viability. Animals exhibited no toxicity in the parameters analyzed in acute oral toxicity, comet assay, and MN tests. Further, EtEx promoted a significant reduction in DNA damage index and DNA damage frequency utilizing the comet assay, while the group treated with AqEx exhibited no marked differences. Thus, data demonstrated that AqEx or EtEx of embiratanha may be considered safe at a dose of 2,000 mg/kg orgally under our experimental conditions tested.

Study on Quality Characteristic of Chebulae Fructus and Its Adulterants and Degradation Pathway of Hydrolyzable Tannins.

Chebulae Fructus (CF) is known as one of the richest sources of hydrolyzable tannins (HTs). In this study, ultra-performance liquid chromatography coupled with a photodiode array detector method was established for simultaneous determination of the 12 common phenolcarboxylic and tannic constituents (PTCs). Using this method, quantitative analysis was accomplished in CF and other four adulterants, including Terminaliae Belliricae Fructus, Phyllanthi Fructus, Chebulae Fructus Immaturus, and Canarii Fructus. Based on a quantitative analysis of the focused compounds, discrimination of CF and other four adulterants was successfully accomplished by hierarchical cluster analysis and principal component analysis. Additionally, the total contents of the 12 compounds that we focused on in this study were unveiled as 148.86 mg/g, 96.14 mg/g, and 18.64 mg/g in exocarp, mesocarp, and endocarp and seed of CF, respectively, and PTCs were witnessed to be the most abundant in the exocarp of CF. Noticeably, the HTs (chebulagic acid, chebulanin acid, chebulinic acid, and punicalagin) were observed to be ultimately degraded to chebulic acid, gallic acid, and ellagic acid during sunlight-drying of the fresh fruits. As a result, our study indicated that CF and its adulterants could be distinguished by the observed 12 PTCs, which were mainly distributed in the exocarp of the fruits. The HTs were prone to degrade into the three simple phenolcarboxylic acids during drying or processing, allowing us to obtain a more comprehensive understanding of the PTCs, with great significance in the improved quality of CF and related products.

Reconnoitring the antioxidant and antibacterial potential of different fruits after tannin acyl hydrolase mediated biotransformation.

Condensed and hydrolyzable tannins are secondary metabolites present in almost every plant part. Tannase enzyme acts on hydrolyzable tannins to produce gallic acid and tannase-mediated end-products with immense therapeutic potential. Seven different fruits with significant presence of hydrolyzable tannin content were selected to check for phenol, tannin, and hydrolyzable tannin contents. Prunus domestica had the maximum phenol content, that is, 85.4 ± 0.207, followed by Syzygium cumini, Fragaria ananassa, Rubus fruticosus, and Psidium guajava. Plum showed the maximum number of hydrolyzable tannins. Fruit extracts were subjected to tannase hydrolysis and their antimicrobial and antioxidant activities were determined. There was a significant increase in the antioxidant abilities of the fruits with Punica granatum extract, displaying the highest decline of 132 units of IC50 followed by F. ananassa hydrolyzable extract, showing a decrease from 224.75 to 119.98 µg/mL. The extracts also depicted a significant increase in antibacterial activity after hydrolysis against Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis, and Staphylococcus aureus with Rubus idaeus aqueous extract observed to be most effective against E. coli. The increase in antioxidant and antibacterial activity can be attributed to the production of tannase-mediated products formed after the biotransformation of hydrolyzable tannins present in the aqueous extracts.

The chromosome-scale reference genome of Rubus chingii Hu provides insight into the biosynthetic pathway of hydrolyzable tannins.

Rubus chingii Hu (Fu-Pen-Zi), a perennial woody plant in the Rosaceae family, is a characteristic traditional Chinese medicinal plant because of its unique pharmacological effects. There are abundant hydrolyzable tannin (HT) components in R. chingii that provide health benefits. Here, an R. chingii chromosome-scale genome and related functional analysis provide insights into the biosynthetic pathway of HTs. In total, sequence data of 231.21 Mb (155 scaffolds with an N50 of 8.2 Mb) were assembled into seven chromosomes with an average length of 31.4 Mb, and 33 130 protein-coding genes were predicted, 89.28% of which were functionally annotated. Evolutionary analysis showed that R. chingii was most closely related to Rubus occidentalis, from which it was predicted to have diverged 22.46 million years ago (Table S8). Comparative genomic analysis showed that there was a tandem gene cluster of UGT, carboxylesterase (CXE) and SCPL genes on chromosome 02 of R. chingii, including 11 CXE, eight UGT, and six SCPL genes, which may be critical for the synthesis of HTs. In vitro enzyme assays indicated that the proteins encoded by the CXE (LG02.4273) and UGT (LG02.4102) genes have tannin hydrolase and gallic acid glycosyltransferase functions, respectively. The genomic sequence of R. chingii will be a valuable resource for comparative genomic analysis within the Rosaceae family and will be useful for understanding the biosynthesis of HTs.

Do Pomegranate Hydrolyzable Tannins and Their Derived Metabolites Provide Relief in Osteoarthritis? Findings from a Scoping Review.

Osteoarthritis (OA) is the most common form of arthritis affecting both the elderly and the middle-aged population. Although various therapeutics have been developed to arrest the structural deterioration of cartilage, the current treatments are limited to delay the progress of OA clinically. Therefore, it is pivotal to study new therapeutic agents for chondroprotection and the prevention of cartilage degeneration. Hydrolyzable tannin (HT)-containing foods aroused considerable interest in recent years for their relevant anti-inflammatory effects. The focus of this scoping review is to provide an overview of the evidence of the therapeutic potential of HTs and their metabolites in preventing or alleviating the course of OA. A broad search of PubMed and Scopus databases on this topic resulted in 156 articles. After the exclusion of reviews and not relevant records, 31 articles were retrieved. Although only some papers did not consider the biotransformation of HTs, most recent studies also have investigated the effect of HT metabolites. Further larger clinical trials, with an in-deep analysis of HT metabolization, are still needed to unravel the potential benefits of these compounds in OA, paving the way towards the development of a dietary strategy for the improvement of pro-inflammatory cytokine-induced chondrocyte dysfunctions and injuries.

Hydrolyzable Tannins in the Management of Th1, Th2 and Th17 Inflammatory-Related Diseases.

Plants rich in hydrolyzable tannins were traditionally used all over the world for a variety of chronic inflammatory disorders, including arthritis, colitis, and dermatitis. However, the knowledge of their immunological targets is still limited though fundamental for their rational use in phytotherapy. The recent advances regarding the pathogenesis of inflammatory-based diseases represent an opportunity to elucidate the pharmacological mechanism of plant-derived metabolites with immunomodulatory activity. This review collects recent articles regarding the role of hydrolyzable tannins and their gut metabolites in Th1, Th2, and Th17 inflammatory responses. In line with the traditional use, rheumatoid arthritis (RA), inflammatory bowel diseases (IBDs), psoriasis, atopic dermatitis (AD), and asthma were the most investigated diseases. A substantial body of in vivo studies suggests that, beside innate response, hydrolyzable tannins may reduce the levels of Th-derived cytokines, including IFN-γ, IL-17, and IL-4, following oral administration. The mode of action is multitarget and may involve the impairment of inflammatory transcription factors (NF-κB, NFAT, STAT), enzymes (MAPKs, COX-2, iNOS), and ion channels. However, their potential impact on pathways with renewed interest for inflammation, such as JAK/STAT, or the modulation of the gut microbiota demands dedicate studies.

Oenothein B in Eucalyptus Leaf Extract Suppresses Fructose Absorption in Caco-2 Cells.

Inhibition of fructose absorption may suppress adiposity and adiposity-related diseases caused by fructose ingestion. Eucalyptus leaf extract (ELE) inhibits intestinal fructose absorption (but not glucose absorption); however, its active compound has not yet been identified. Therefore, we evaluated the inhibitory activity of ELE obtained from Eucalyptus globulus using an intestinal fructose permeation assay with the human intestinal epithelial cell line Caco-2. The luminal sides of a cell monolayer model cultured on membrane filters were exposed to fructose with or without the ELE. Cellular fructose permeation was evaluated by measuring the fructose concentration in the medium on the basolateral side. ELE inhibited 65% of fructose absorption at a final concentration of 1 mg/mL. Oenothein B isolated from the ELE strongly inhibited fructose absorption; the inhibition rate was 63% at a final concentration of 5 µg/mL. Oenothein B did not affect glucose absorption. In contrast, the other major constituents (i.e., gallic acid and ellagic acid) showed little fructose-inhibitory activity. To our knowledge, this is the first report that oenothein B in ELE strongly inhibits fructose absorption in vitro. ELE containing oenothein B can prevent and ameliorate obesity and other diseases caused by dietary fructose consumption.

Bacterial tannases: classification and biochemical properties.

Tannin acyl hydrolases, also known as tannases, are a group of enzymes critical for the transformation of tannins. The study of these enzymes, which initially evolved in different organisms to detoxify and/or use these plant metabolites, has nowadays become relevant in microbial enzymology research due to their relevant role in food tannin transformation. Microorganisms, particularly bacteria, are major sources of tannase. Cloning and heterologous expression of bacterial tannase genes and structural studies have been performed in the last few years. However, a systematic compilation of the information related to all recombinant tannases, their classification, and characteristics is missing. In this review, we explore the diversity of heterologously produced bacterial tannases, describing their substrate specificity and biochemical characterization. Moreover, a new classification based on sequence similarity analysis is proposed. Finally, putative tannases have been identified in silico for each group of tannases taking advantage of the use of the "tannase" distinctive features previously proposed.

Induced plant defenses, host-pathogen interactions, and forest insect outbreaks.

Cyclic outbreaks of defoliating insects devastate forests, but their causes are poorly understood. Outbreak cycles are often assumed to be driven by density-dependent mortality due to natural enemies, because pathogens and predators cause high mortality and because natural-enemy models reproduce fluctuations in defoliation data. The role of induced defenses is in contrast often dismissed, because toxic effects of defenses are often weak and because induced-defense models explain defoliation data no better than natural-enemy models. Natural-enemy models, however, fail to explain gypsy moth outbreaks in North America, in which outbreaks in forests with a higher percentage of oaks have alternated between severe and mild, whereas outbreaks in forests with a lower percentage of oaks have been uniformly moderate. Here we show that this pattern can be explained by an interaction between induced defenses and a natural enemy. We experimentally induced hydrolyzable-tannin defenses in red oak, to show that induction reduces variability in a gypsy moth's risk of baculovirus infection. Because this effect can modulate outbreak severity and because oaks are the only genus of gypsy moth host tree that can be induced, we extended a natural-enemy model to allow for spatial variability in inducibility. Our model shows alternating outbreaks in forests with a high frequency of oaks, and uniform outbreaks in forests with a low frequency of oaks, matching the data. The complexity of this effect suggests that detecting effects of induced defenses on defoliator cycles requires a combination of experiments and models.

Genome-wide identification of TCP transcription factors and their potential roles in hydrolyzable tannin production in Quercus variabilis cupule.

Hydrolyzable tannins (HTs) have garnered significant attention due to their proven beneficial effects in the clinical treatment of various diseases. The cupule of Chinese cork oak (Quercus variabilis Blume) has been used as raw material of traditional medicine for centuries for its high content of HTs. Previous studies have identified UGT84A13 as a key enzyme in the HT biosynthesis pathway in Q. variabilis, but the transcriptional regulation network of UGT84A13 remains obscure. Here, we performed a comprehensive genome-wide identification of the TCP transcription factors in Q. variabilis, elucidating their molecular evolution and gene structure. Gene expression analysis showed that TCP3 from the CIN subfamily and TCP6 from the PCF subfamily were co-expressed with UGT84A13 in cupule. Further functional characterization using dual-luciferase assays confirmed that TCP3, rather than TCP6, played a role in the transcriptional regulation of UGT84A13, thus promoting HT biosynthesis in the cupule of Q. variabilis. Our work identified TCP family members in Q. variabilis for the first time, and provided novel insights into the transcriptional regulatory network of UGT84A13 and HT biosynthesis in Q. variabilis, explaining the reason why the cupule enriches HTs that could be used for traditional medicine.

Influence of fecal fermentation on the anthelmintic activity of proanthocyanidins and ellagitannins against human intestinal nematodes and Caenorhabditis elegans.

Some tannin-rich plants such as Combretum mucronatum and Phyllanthus urinaria are widely used in Africa for the control of parasitic nematodes in both humans and livestock. Tannins have been recognized as an alternative source of anthelmintic therapies, and hence, recent studies have focused on both the hydrolyzable and condensed tannins. These groups of compounds, however, have poor oral bioavailability and are metabolized by gut microbiota into lower molecular weight compounds. The role of these metabolites in the anthelmintic activities of tannins has not been explored yet. This study investigated the effects of fecal metabolism on the anthelmintic potential of procyanidin C1 (PC1) and geraniin and the tannin-enriched extracts of C. mucronatum (CML) and P. urinaria (PUH), which contain these compounds, respectively. Metabolites were formed by anaerobic fermentation of the test compounds and extracts in a fresh human fecal suspension for 0 h, 4 h, and 24 h. Lyophilized samples were tested in vitro against hookworm larvae and whipworm (Trichuris trichiura) larvae obtained from naturally infected human populations in Pru West District, Bono East Region, Ghana, and against the wildtype strain of Caenorhabditis elegans (L4). Both extracts and compounds in the undegraded state exhibited concentration-dependent inhibition of the three nematodes. Their activity, however, significantly decreased upon fecal metabolism. Without fermentation, the proanthocyanidin-rich CML extract was lethal against hookworm L3 (LC50 = 343.5 µg/mL, 95% confidence interval (CI) = 267.5-445.4), T. trichiura L1 (LC50 = 230.1 µg/mL, CI = 198.9-271.2), and C. elegans (LC50 = 1468.1 µg/mL, CI = 990.3-1946.5). PUH, from which the ellagitannin geraniin was isolated, exhibited anthelmintic effects in the unfermented form with LC50 of 300.8 µg/mL (CI = 245.1-374.8) against hookworm L3 and LC50 of 331.6 µg/mL (CI = 290.3-382.5) against T. trichiura L1, but it showed no significant activity against C. elegans L4 larvae at the tested concentrations. Similarly, both compounds, procyanidin C1 and geraniin, lost their activity when metabolized in fecal matter. The activity of geraniin at a concentration of 170 µg/mL against C. elegans significantly declined from 30.4% ± 1.8% to 14.5% ± 1.5% when metabolized for 4 h, whereas that of PC1 decreased from 32.4% ± 2.3% to 8.9% ± 0.9% with similar treatment. There was no significant difference between the anthelmintic actions of metabolites from the structurally different tannin groups. The outcome of this study revealed that the intact bulky structure of tannins (hydrolyzable or condensed) may be required for their anthelmintic action. The fermented products from the gut may not directly contribute toward the inhibition of the larvae of soil-transmitted helminths.

The Impact of Mineral and Energy Supplementation and Phytogenic Compounds on Rumen Microbial Diversity and Nitrogen Utilization in Grazing Beef Cattle.

The objective of this study was to evaluate the effect of the addition of a phytogenic compound blend (PHA) containing hydrolyzable tannins, carvacrol, and cinnamaldehyde oil to mineral salt or energy supplementation on the rumen microbiota and nitrogen metabolism of grazing Nellore cattle. Eight castrated Nellore steers were distributed in a double-Latin-square 4 × 4 design, with a 2 × 2 factorial arrangement (two types of supplements with or without the addition of the PHA), as follows: energy supplement without the PHA addition (EW); energy supplement with the PHA addition (EPHA); mineral supplement without the addition of the PHA (MW); mineral supplement with the PHA addition (MPHA). Steers that received supplements with the PHA have a lower ruminal proportion of valerate (with the PHA, 1.06%; without the PHA, 1.15%), a lower ruminal abundance of Verrucomicrobia, and a tendency for lower DM digestibility (with the PHA, 62.8%; without the PHA, 64.8%). Energy supplements allowed for higher ammonia concentrations (+2.28 mg of NH3-N/dL), increased the propionate proportion (+0.29% of total VFA), and had a higher ruminal abundance of Proteobacteria and Spirochaetae phyla in the rumen. The PHA addition in the supplement did not improve nitrogen retention, reduced the ruminal proportion of valerate, and had a negative impact on both the total dry-matter digestibility and the abundance of several ruminal bacterial groups belonging to the Firmicutes and Verrucomicrobia phyla.

New Hydrolyzable Tannin with Potent Antioxidant and α-Glucosidase Inhibitory Activity from Black Tea Produced from Camellia taliensis.

Camellia taliensis (W. W. Smith) Melchior, belonging to the genus Camellia sect. Thea., is mainly distributed from northern Myanmar to western and southwestern Yunnan province of China, and its leaves have been used to make various teas by the locals of its growing regions. The chemical constituents of C. taliensis are significantly related to those of cultivated tea plants, C. sinensis and C. sinensis var. assamica. The HPLC-ESI-MS analysis of black tea prepared from the leaves of C. taliensis showed a rich existence of polyphenols. Further comprehensive chemical study led to the separation and recognition of 32 compounds (1-32), including one new hydrolyzable tannin, 1-O-galloyl-4,6-tetrahydroxydibenzofurandicarboxyl-ß-D-glucopyranose (1), and one new natural product (24). The known compounds referred to seven hydrolyzable tannins (2-8), 10 flavonols and glycosides (9-18), and 14 simple phenolics (19-32). Their structures were elucidated by comprehensive spectroscopic analyses. Among them, 20 compounds (2, 3, 6, 7, 8, 15, 17, 18, 20-22, 24-32) were isolated from black tea for the first time. Most isolates displayed obvious antioxidant activities on DPPH and ABTS+ assays, and the hydrolyzable tannins 1, 3-5, 7, and 8 exhibited stronger inhibitory activities on α-glycosidase than quercetin and acarbose (IC50 = 5.75 and 223.30 µM, respectively), with IC50 values ranging from 0.67 to 2.01 µM.

Possible Beneficial Effects of Hydrolyzable Tannins Deriving from Castanea sativa L. in Internal Medicine.

Hydrolyzable tannins (HTs) deriving from chestnuts have demonstrated, through numerous studies, the ability to exert multiple beneficial effects, including antioxidant and antimicrobial effects, on the lipid metabolism and cancer cells. The latter effect is very fascinating, since different polyphenols deriving from chestnuts were able to synergistically induce the inhibition of cancerous cells through multiple pathways. Moreover, the main mechanisms by which tannins induce antioxidant functions include: the reduction in oxidative stress, the ability to scavenge free radicals, and the modulation of specific enzymes, such as superoxide dismutase. HTs have also been shown to exert significant antimicrobial activity by suppressing microbial growth. The actions on the lipid metabolism are several, among which is the inhibition of lipid accumulation. Thus, tannins seem to induce a cardioprotective effect. In fact, through various mechanisms, such as the relaxation of the vascular smooth muscle, HTs were proven to be efficient against arterial hypertension. Therefore, the great number of studies in this field prove the growing interest on the utilization of natural bioactive compounds, such as HTs deriving from natural sources or obtained by circular economy models, as potential nutraceuticals or adjuvants therapies.

Valorization of Prickly Pear Peel Residues (Opuntia ficus-indica) Using Solid-State Fermentation.

Prickly pear peel (Opuntia ficus-indica) residues can be used as a substrate in solid-state fermentation to obtain bioactive compounds. The kinetic growth of some Aspergillus strains was evaluated. A Box-Hunter and Hunter design to evaluate the independent factors was used. These factors were temperature (°C), inoculum (spores/g), humidity (%), pH, NaNO3 (g/L), MgSO4 (g/L), KCl (g/L), and KH2PO4 (g/L). The response factors were the amount of hydrolyzable and condensed tannins. The antioxidant and antimicrobial activity of fermentation extracts was evaluated. Aspergillus niger strains GH1 and HT3 were the best for accumulating tannins. The humidity, inoculum, and temperature affect the release of hydrolyzable and condensed tannins. Treatment 13 (low values for temperature, inoculum, NaNO3, MgSO4; and high values for humidity, pH, KCl, KH2PO4) resulted in 32.9 mg/g of condensed tannins being obtained; while treatment 16 (high values for all the factors evaluated) resulted in 3.5 mg/g of hydrolyzable tannins being obtained. In addition, the fermented extracts showed higher antioxidant activity compared to the unfermented extracts. Treatments 13 and 16 showed low inhibition of E. coli, Alternaria sp., and Botrytis spp. The solid-state fermentation process involving prickly pear peel residues favors the accumulation of condensed and hydrolyzable tannins, with antioxidant and antifungal activity.

Solid-State Fermentation of Sorghum by Aspergillus oryzae and Aspergillus niger: Effects on Tannin Content, Phenolic Profile, and Antioxidant Activity.

Sorghum contains antioxidants such as tannins. However, these are considered antinutritional factors since they are responsible for the low digestibility of proteins and carbohydrates. Nevertheless, these can be extracted by solid-state fermentation (SSF). Therefore, this study aimed to evaluate the effects of SSF from Aspergillus oryzae and Aspergillus niger Aa210 on the tannin contents, phenolic profiles determined by HPLC-MS, and antioxidant activities (ABTS, DPPH, and FRAP) of two genotypes of sorghum. The results showed that with SSF by A. niger Aa210, a higher tannin content was obtained, with yields of 70-84% in hydrolyzable tannins (HT) and 33-49% in condensed tannins (CT), while with SSF by A. oryzae the content of HT decreased by 2-3% and that of CT decreased by 6-23%. The extracts fermented by A. niger at 72 and 84 h exhibited a higher antioxidant activity. In the extracts, 21 polyphenols were identified, such as procyanidins, (+)-catechin, (-)-epicatechin, scutellarein, arbutin, and eriodictyol, among others. Therefore, SSF by A. niger was an efficient process for the release of phenolic compounds that can be used as antioxidants in different food products. It is also possible to improve the bioavailability of nutrients in sorghum through SSF. However, more studies are required.

Optimization of Deep Eutectic Solvent Extraction of Phenolic Acids and Tannins from Alchemilla vulgaris L.

Alchemilla vulgaris L. is a good source of antioxidant components with an emphasis on phenolic acids and tannins. In this study, gallic acid, ellagic acid, and hydrolyzable tannins (HT) were extracted from this plant with different deep eutectic solvents (DESs), varying the amount of added H2O, temperature and extraction time. Seventeen DESs (n = 3) were used for the extraction, of which choline chloride:urea (1:2) proved to be the most suitable. The selection of the best solvent was followed by the examination of the influence of the extraction type and parameters using response surface methodology (RSM). Gallic acid content was in the range of 0.00-1.89 µg mg-1, ellagic acid content was 0.00-12.76 µg mg-1 and hydrolyzable tannin (HT) content was 3.06-181.26 µgTAE mg-1, depending on the used technique and the extraction conditions. According to the results, extraction by stirring and heating was the most suitable since the highest amounts of gallic acid, ellagic acid, and HT were extracted, and the obtained optimal values using response surface methodology (RSM) are confirmed by experimentally obtained values.

Diospyros villosa Root Monographic Quality Studies.

Diospyros villosa L. (De Winter) (Ebenaceae) is a shrub whose root (DVR) is used as a toothbrush and to treat oral infections in Mozambique. The present work aims at establishing monographic quality criteria to allow the sustainable and safe development of pharmaceutical preparations with this herbal drug. This includes setting botanical (qualitative and quantitative) and chemical identification parameters, purity tests (loss on drying and total ash), quantifying the major classes of constituents identified, and particle size characterization of the powdered drug. DVR samples are cylindrical and microscopically characterized by: a periderm, with six layers of flattened phellem cells, with slightly thickened walls and few layers of phelloderm; cortical parenchyma with brachysclereids with a short, roughly isodiametric form (13.82-442.14 µm2 × 103), surrounded by a ring of prismatic calcium oxalate crystals; uniseriate medullary rays and prominent vessels of the xylem with single or double shape; numerous single and clustered starch grains, within the cortical parenchyma, medullar parenchyma, and ray cells. Polyphenols, mainly hydrolyzable tannins (212.29 ± 0.005 mg gallic acid equivalent/g of dried DVR), are the main marker class of constituents. Furthermore, the average diameter of the particles of the powder, 0.255 mm, allows its classification as a fine powder.