Quality Studies on Cynometra iripa Leaf and Bark as Herbal Medicines.

Cynometra iripa Kostel. is a Fabaceae species of mangrove used in traditional Ayurvedic medicine for treating inflammatory conditions. The present study aims to establish monographic botanical and chemical quality criteria for C. iripa leaf and bark as herbal substances and to evaluate their in vitro antioxidant potential. Macroscopic and microscopic qualitative and quantitative analyses, chemical LC-UV/DAD-ESI/MS profiling, and the quantification of key chemical classes were performed. Antioxidant activity was evaluated by DPPH and FRAP assays. Macroscopically, the leaf is asymmetrical with an emarginated apex and cuneate base. Microscopically, it shows features such as two-layered adaxial palisade parenchyma, vascular bundles surrounded by 3-6 layers of sclerenchyma, prismatic calcium oxalate crystals (5.89 ± 1.32 µm) along the fibers, paracytic stomata only on the abaxial epidermis (stomatal index-20.15), and non-glandular trichomes only on petiolules. The microscopic features of the bark include a broad cortex with large lignified sclereids, prismatic calcium oxalate crystals (8.24 ± 1.57 µm), and secondary phloem with distinct 2-5 seriated medullary rays without crystals. Chemical profile analysis revealed that phenolic derivatives, mainly condensed tannins and flavonoids, are the main classes identified. A total of 22 marker compounds were tentatively identified in both plant parts. The major compounds identified in the leaf were quercetin-3-O-glucoside and taxifolin pentoside and in the bark were B-type dimeric proanthocyanidins and taxifolin 3-O-rhamnoside. The total phenolics content was higher in the leaf (1521 ± 4.71 mg GAE/g dry weight), while the total flavonoids and condensed tannins content were higher in the bark (82 ± 0.58 mg CE/g and 1021 ± 5.51 mg CCE/g dry weight, respectively). A total of 70% of the hydroethanolic extracts of leaf and bark showed higher antioxidant activity than the ascorbic acid and concentration-dependent scavenging activity in the DPPH assay (IC50 23.95 ± 0.93 and 23.63 ± 1.37 µg/mL, respectively). A positive and statistically significant (p < 0.05) correlation between the phenol content and antioxidant activity was found. The results obtained will provide important clues for the quality control criteria of C. iripa leaf and bark, as well as for the knowledge of their pharmacological potential as possible anti-inflammatory agents with antioxidant activity.

An Integrated Approach to the Anti-Inflammatory, Antioxidant, and Genotoxic Potential of Portuguese Traditional Preparations from the Bark of Anacardium occidentale L.

Anacardium occidentale L. stem bark Traditional Herbal Preparations (AoBTHPs) are widely used in traditional medicine to treat inflammatory conditions, such as diabetes. The present study aims to evaluate the anti-inflammatory, antioxidant, and genotoxic potential of red and white Portuguese AoBTHPs. Using a carrageenan-induced rat paw edema model, a significant anti-edema effect was observed for all tested doses of white AoBTHP (40.2, 71.5, and 127.0 mg/kg) and the two highest doses of red AoB THP (71.5 and 127.0 mg/kg). The anti-edema effect of red AoBTHP's highest dose was much more effective than indomethacin 10 mg/kg, Trolox 30 mg/kg, and Tempol 30 mg/kg. In DPPH, FRAP, and TAC using the phosphomolybdenum method, both types of AoBTHPs showed similar antioxidant activity and no genotoxicity up to 5000 µg/plate in the Ames test. The LC-UV/DAD-ESI/MS fingerprint allowed the identification of gallic and protocatechuic acids as the two main marker compounds and the presence of catechin, epicatechin, epigallocatechin gallate, and ellagic acid in both AoBTHPs. The obtained results support the validation of red and white AoB and their THPs as anti-inflammatory agents and contribute to the possible development of promising new therapeutic options to treat inflammatory conditions.

Quality, Safety and Biological Studies on Campylanthus glaber Aerial Parts.

In Cabo Verde, several endemic species are used in traditional medicine. However, no scientific studies have been conducted on the quality, efficacy, and safety of most of these plants. This study focused on establishing the botanical and chemical identification parameters required for a quality monograph of Campylanthus glaber Benth. aerial parts, a medicinal plant of Cabo Verde traditionally used to treat fever and muscular pain. In addition, in vitro antioxidant and antihyperglycemic activity, cytotoxicity, and genotoxicity were assessed for this medicinal plant. Optical microscopy, LC/UV-DAD-ESI/MS, and colorimetric assays were used for botanical, chemical, and biological studies, respectively. Cytotoxicity was assessed by the MTT assay with HepG2 cells, and genotoxicity by the Ames test. Microscopically, the xeromorphic leaf of C. glaber presents a thick cuticle (13.6-25.5 µm), thick-walled epidermal cells, anomocytic-type stomata, glandular trichomes (stalk length = 49.4-120.8 µm), and idioblasts containing calcium oxalate microcrystals. The chemical screening of aqueous and hydroethanolic extracts of this medicinal plant revealed the presence of organic acids, iridoids, phenylethanoids, and flavonoids as the main classes of marker compounds, with malic acid, citric acid, and verbascoside being the main marker compounds identified. Both extracts showed similar LC/UV-DAD/ESI-MS qualitative profiles and DPPH radical scavenger activity (IC50 = 130.9 ± 1.4; 134.3 ± 3.1 µg/mL). The hydroethanolic extract inhibited both α-amylase and α-glucosidase enzymes in a dose-dependent manner. Both extracts showed no cytotoxicity (up to 1000 µg/mL) by the MTT assay and no genotoxic potential with or without metabolic activation up to 5 mg /plate. The results obtained are an important contribution to the monographic quality assessment of C. glaber aerial parts and suggest that this medicinal plant may be safe and potentially used as an herbal drug raw material for pharmaceutical purposes.

Bioguided Identification of Active Antimicrobial Compounds from Asphodelus bento-rainhae and Asphodelus macrocarpus Root Tubers.

Root tubers of Asphodelus bento-rainhae subsp. bento-rainhae (AbR), a vulnerable endemic species, and Asphodelus macrocarpus subsp. macrocarpus (AmR) have traditionally been used in Portugal to treat inflammatory and infectious skin disorders. The present study aims to evaluate the in vitro antimicrobial activity of crude 70% and 96% hydroethanolic extracts of both medicinal plants, specifically against multidrug-resistant skin-related pathogens, to identify the involved marker secondary metabolites and also to assess the pre-clinical toxicity of these medicinal plant extracts. Bioguided fractionation of the 70% hydroethanolic extracts of both species using solvents of increasing polarity, namely diethyl ether (DEE: AbR-1, AmR-1), ethyl acetate (AbR-2, AmR-2) and aqueous (AbR-3, AmR-3) fractions, enabled the identification of the DEE fractions as the most active against all the tested Gram-positive microorganisms (MIC: 16 to 1000 µg/mL). Furthermore, phytochemical analyses using TLC and LC-UV/DAD-ESI/MS techniques revealed the presence of anthracene derivatives as the main constituents of DEE fractions, and five known compounds, namely 7'-(chrysophanol-4-yl)-chrysophanol-10'-C-beta-D-xylopyranosyl-anthrone (p), 10,7'-bichrysophanol (q), chrysophanol (r), 10-(chrysophanol-7'-yl)-10-hydroxychrysophanol-9-anthrone (s) and asphodelin (t), were identified as the main marker compounds. All these compounds showed high antimicrobial activity, particularly against Staphylococcus epidermidis (MIC: 3.2 to 100 µg/mL). Importantly, no cytotoxicity against HepG2 and HaCaT cells (up to 125 µg/mL) for crude extracts of both species and genotoxicity (up to 5000 µg/mL, with and without metabolic activation) for AbR 96% hydroethanolic extract was detected using the MTT and Ames tests, respectively. Overall, the obtained results contribute to the concrete validation of the use of these medicinal plants as potential sources of antimicrobial agents in the treatment of skin diseases.

Vernonia britteniana Root Phytochemical Studies, In Vitro Cercaricidal Activity on the Larval Stage of Schistosoma mansoni and Antioxidant Activities.

Vernonia britteniana Hiern. (Asteraceae) is a medicinal plant used in traditional Angolan medicine against schistosomiasis. Our study aimed to investigate the phytochemical composition and the cercaricidal and antioxidant activities in vitro of a traditional herbal preparation (Water-Vbr) and a 70% hydroethanolic extract (EtOH70%-Vbr) prepared with this medicinal plant. The activity of the extracts against Schistosoma mansoni cercariae was assessed at different extract concentrations (500, 438, and 125 µg/mL) and at different time intervals, and the phytochemical profiles were obtained by LC-UV-ESI/MS-MS. In addition, the major chemical classes of the identified metabolites were quantified by colorimetry, and the antioxidant potential was assessed using the DPPH and FRAP methods. After 30 min, 100% cercarial mortality was observed at a concentration of 500 µg/mL after exposure, and after 120 min, an LC50 of 438 µg/mL was observed for both extracts. Phenolic acid derivatives (chlorogenic acid, caffeic acid; 3,4-di-O-caffeoylquinic acid; 3,5-di-O-caffeoylquinic acid; and 4,5-di-O-caffeoylquinic acid) and triterpenoids (stigmastane-type steroidal saponins; vernoamyoside D and vernonioside D1; vernoamyoside B; and vernoniamyoside A and C) were identified as the main secondary metabolites. The Water-Vbr extract showed the highest antioxidant activity-DPPH: IC50 = 1.769 ± 0.049 µg/mL; FRAP: mean = 320.80 ± 5.1325 µgAAE/g.

Diversity and biological activities of medicinal plants of Santiago island (Cabo Verde).

Plants continue to constitute key elements of medical practice in West African countries. The Cabo Verde archipelago hosts a great diversity of medicinal plants and local markets are considered important sites for trading plants harvested by rural communities. This study has two main goals: (i) to assess the medicinal uses of native species in Santiago, the biggest island of the archipelago, and (ii) to evaluate the antioxidant, antimicrobial and antidiabetic/antihyperglycemic activities of two native trees (Tamarix senegalensis and Sideroxylon marginatum) used in traditional medicine and traded in local markets. Our results revealed that on Santiago Island, 24 native plants are used in traditional medicine. The main uses of these species (e.g., forage, timber, food and fibres), their medicinal applications, the plant parts used, their mode of administration and conservation status are presented here for the first time. Moreover, the pharmacological characterization of two native tree species revealed that hydroethanolic extracts were richer in phenolic compounds and more active than their aqueous counterparts. All the studied extracts revealed significant antioxidant properties (DPPH and FRAP assays) and were generally moderately active against Gram-positive bacteria. All the extracts inhibited the activities of the carbohydrate digestive enzymes α-glucosidase and α-amylase in a dose-dependent manner. For α-glucosidase, the detected inhibitory activity (IC50 values from 2.0 ± 0.2 µg/mL to 9.9 ± 1.2 µg/mL) was significantly higher than that of acarbose, suggesting that extracts of both species can delay glucose absorption, thereby assisting in slowing down the progression of diabetes. Our findings highlight the crucial importance that medicinal plants have for the Cabo Verdean population, while also raising awareness on the need for sustainable use and conservation of native flora, and of tree species traded in local markets in particular.

Identification of Marker Compounds and In Vitro Toxicity Evaluation of Two Portuguese Asphodelus Leaf Extracts.

The leaves of Asphodelus bento-rainhae subsp. bento-rainhae, an endemic Portuguese species, and Asphodelus macrocarpus subsp. macrocarpus have been used as food, and traditionally as medicine, for treating ulcers, urinary tract, and inflammatory disorders. The present study aims to establish the phytochemical profile of the main secondary metabolites, together with the antimicrobial, antioxidant and toxicity assessments of both Asphodelus leaf 70% ethanol extracts. Phytochemical screenings were conducted by the TLC and LC-UV/DAD-ESI/MS chromatographic technique, and quantification of the leading chemical classes was performed by spectrophotometric methods. Liquid-liquid partitions of crude extracts were obtained using ethyl ether, ethyl acetate, and water. For in vitro evaluations of antimicrobial activity, the broth microdilution method, and for the antioxidant activity, the FRAP and DPPH methods were used. Genotoxicity and cytotoxicity were assessed by Ames and MTT tests, respectively. Twelve known compounds including neochlorogenic acid, chlorogenic acid, caffeic acid, isoorientin, p-coumaric acid, isovitexin, ferulic acid, luteolin, aloe-emodin, diosmetin, chrysophanol, and ß-sitosterol were identified as the main marker compounds, and terpenoids and condensed tannins were found to be the major class of secondary metabolites of both medicinal plants. The ethyl ether fractions demonstrated the highest antibacterial activity against all the Gram-positive microorganisms, (MIC value of 62 to 1000 µg/mL), with aloe-emodin as one of the main marker compounds highly active against Staphylococcus epidermidis (MIC value of 0.8 to 1.6 µg/mL). Ethyl acetate fractions exhibited the highest antioxidant activity (IC50 of 800 to 1200 µg/mL, respectively). No cytotoxicity (up to 1000 µg/mL) or genotoxicity/mutagenicity (up to 5 mg/plate, with/without metabolic activation) were detected. The obtained results contribute to the knowledge of the value and safety of the studied species as herbal medicines.

Monographic Quality Parameters and Genotoxicity Assessment of Asphodelus bento-rainhae and Asphodelus macrocarpus Root Tubers as Herbal Medicines.

Root tubers of Asphodelus bento-rainhae subsp. bento-rainhae (AbR), an endemic species with relevant interest due to conservation concerns, and Asphodelus macrocarpus subsp. macrocarpus (AmR) have been traditionally used for culinary and medicinal purposes, mainly associated with skin infection and inflammation. The present study aims to establish the quality control criteria for the proper characterization of dried root tubers of both species as herbal substances, together with their preclinical safety assessments. Botanical identification using macroscopic and microscopic techniques and phytochemical evaluation/quantification of the main classes of marker secondary metabolites, including phenolic compounds (flavonoid, anthraquinone, condensed and hydrolysable tannin) and terpenoids were performed. Additionally, in vitro genotoxicity/mutagenicity was evaluated by Ames test. Evident morphological differences in the development of tubercles (3.5 × 1 cm in AbR and 8.7 × 1.4 cm in AmR) and microscopicly in the arrangements and characteristics of the vascular cylinder (metaxylem and protoxylems) were found. Anatomical similarities such as multiple-layered epidermis (velamen) and the cortex area with thin-walled idioblasts (134 ± 2.9 µm and 150 ± 27.6 µm) containing raphide crystals (37.2 ± 14.2 µm and 87.7 ± 15.3 µm) were observed between AbR and AmR, respectively. Terpenoids (173.88 ± 29.82 and 180.55 ± 10.57 mg OAE/g dried weight) and condensed tannins (128.64 ± 14.05 and 108.35 ± 20.37 mg CAE/g dried weight) were found to be the main class of marker secondary metabolites of AbR and AmR extracts, respectively. No genotoxicity (up to 5 mg/plate, without metabolic activation) was detected in these medicinal plants' tested extracts. The obtained results will contribute to the knowledge of the value of the Portuguese flora and their future commercial cultivation utilization as raw materials for industrial and pharmaceutical use.

Contribution to the Preclinical Safety Assessment of Lannea velutina and Sorindeia juglandifolia Leaves.

Dried leaves of Lannea velutina A. Rich. and Sorindeia juglandifolia (A. Rich.) Planch. ex Oliv. (family Anacardiaceae) are used in African traditional medicine. Although these medicinal plants have widespread use in the treatment of inflammatory diseases, there is no scientific data concerning their preclinical or clinical safety. This work aimed to investigate the phytochemical properties of the leaves of both species using HPLC-UV/DAD, as well as the in vivo oral repeated-dose toxicity of 70% hydroethanolic leaf extract of S. juglandifolia and the in vitro genotoxicity of 70% hydroethanolic leaf extracts of L. velutina and S. juglandifolia. Clinical signs of toxicity, body weight variations, and changes in food consumption, mortality, and blood biochemical parameters were monitored. Genotoxicity was assessed using the bacterial reverse mutation assay (Ames test) with and without metabolic activation, according to OECD guidelines. The obtained results showed the presence of gallic acid and anacardic acid as the main marker constituents in both species. No significant changes in general body weight or food intake were observed; small significant changes with no critical relevance were observed in the blood biochemistry of animals treated with S. juglandifolia hydroethanolic extract (50, 400, and 1000 mg/kg body weight) compared to those in the control group. No genotoxicity was observed in the bacterial reverse mutation assay with S. juglandifolia and L. velutina extracts (up to 5 mg/plate). The safety data obtained in vivo and lack of genotoxic potential in vitro points to the safe medicinal use of S. juglandifolia and L. velutina extracts.

Influence of the in vitro gastrointestinal digestion on the antioxidant activity of Artemisia gorgonum Webb and Hyptis pectinata (L.) Poit. infusions from Cape Verde.

The phenolic profile and antioxidant activity of Cape Verde's Artemisia gorgonum Webb and Hyptis pectinata (L.) Poit. infusions before and after in vitro simulation of the gastrointestinal digestion were determined. The LC-UV/DAD fingerprinting analysis allowed the identification of 3-O-caffeoylquinic acid, chlorogenic acid, 3,5 dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid and other caffeoylquinic acids derivatives on A. gorgonum infusion, and of caffeoylquinic acid and quercetin derivatives on H. pectinata infusion. Despite some decrease in the chromatographic area of several peaks, no relevant qualitative alterations on the chromatographic profile were observed between the digested and undigested herbal infusions. Results obtained showed a decrease on the antioxidant capacity of both tested herbal infusions after the in vitro digestion. This decrease was more pronounced for H. pectinata than for A. gorgonum and was also more pronounced regarding the radical scavenging capacity than regarding the reducing capacity. After complete digestion the superoxide anion and the DPPH-radical scavenging capacities decreased ≈ 43 and 75% for H. pectinata and ≈ 31 and 70% for A. gorgonum. Despite the observed differences before and after simulated gastrointestinal digestion, both infusions still had antioxidant activity at the end of this process. Thus, the antioxidant potential of A. gorgonum and H. pectinata infusions from Cape Verde, prepared as traditionally used, seems to be kept in some extend throughout the digestive system.