Development of AKR1B10 inhibitors from Ajuga nipponensis based on diseases and targets.

Ten compounds (1-10) including one new neoclerodane diterpenoid (1) and nine known compounds were isolated from the whole plants of Ajuga nipponensis. Their structures were established by performing detailed analysis of NMR, the structure of 1 was determined using HRESIMS, 1D and 2D NMR, UV, and IR. Compounds 1 and 4-10 were isolated from Ajuga nipponensis for the first time. And it was the first time to report compounds 9 and 10 as natural products. Based on network pharmacology methods, 45 key targets were selected, which were compounds mapping to diseases. And compounds 2, 3, 7, and a (ajugacumbin B) exhibited excellent AKR1B10 inhibitory activities, with IC50 values of 53.05 ± 0.75, 87.22 ± 0.85, 61.85 ± 0.66, and 85.19±1.02 nM respectively, with Epalrestat used as the positive control (82.09 ± 1.62 nM). Additionally, the interaction between active compounds and AKR1B10 had been discussed according to the molecular docking results. Ultimately, the analysis of GO and KEGG enrichment indicated that the key signaling pathway of the active compounds may be related to prostate cancer. Our study results demonstrate the hypoglycemic and anti-tumor properties of A. nipponensis for the first time, and provide a comprehensive understanding of its application in traditional medicine. Furthermore, this article establishes a reference for further research on the optimized experimental design of novel AKR1B10 inhibitors.

Green synthesised AuNps using Ajuga Bracteosa extract and AuNps-Free supernatant exhibited equivalent antibacterial and anticancerous efficacies.

The current study is designed to synthesize gold nanoparticles using Ajuga bracteosa extract, which is a highly known medicinal herb found in the northern Himalayas. The synthesized gold nanoparticles were initially characterized by UV-Vis spectrophotometer, SEM, FTIR, pXRD, and, GC-MS. Antibacterial efficacy of A. bracteosa extract, AuNps, and AuNps-free supernatant activity was checked against highly pathogenic clinical isolates of Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa via agar well diffusion method, assuming that supernatant might have active compounds. The Nps-free supernatant showed the maximum antibacterial activity against E. coli (20.8±0.3 mm), Staphylococcus aureus (16.5±0.5), and Pseudomonas aeruginosa (13±0.6). While green synthesized AuNps showed effective antibacterial activity (Escherichia coli (16.4±0.3mm), Staphylococcus aureus (15.05±0.5mm), and Pseudomonas aeruginosa (11.07±0.6mm)) which was high compared to A. bracteosa extract. Anticancer activity was assessed by MTT assay on U87 and HEK293 cell lines. Aj-AuNps have an antigrowth effect on both the cell lines however Aj-AuNps-free supernatant which was also evaluated along with the Aj-AuNps, showed high toxicity toward HEK293 cell line compared to U87. Further, the GC-MS analysis of supernatant showed the presence of resultant toxic compounds after the reduction of gold salt, which include Trichloromethane, Propanoic acid, 2-methyl-, methyl ester, Methyl isovalerate, Pentanoic acid, 2-hydroxy-4-methyl-, Benzene-propanoic acid, and alpha-hydroxy. Based on the observation small molecular weight ligands of Ajuga bracteosa were analyzed in-silico for their binding efficacy towards selected membrane proteins of our target pathogens. RMSD is also calculated for the best docked protein ligand pose. The results revealed that among all listed ligands, Ergosterol and Decacetylajugrin IV have high virtuous binding affinities towards the membrane proteins of targeted pathogens. The current findings revealed that the Aj-AuNps are good antibacterial as well as anticancerous agents while the Nps-free supernatant is also exceedingly effective against resistant pathogens and cancer cell lines.

Sustainable Production of Ajuga Bioactive Metabolites Using Cell Culture Technologies: A Review.

The genus Ajuga (Lamiaceae) is rich in medicinally important species with biological activities ranging from anti-inflammatory, antitumor, neuroprotective, and antidiabetic to antibacterial, antiviral, cytotoxic, and insecticidal effects. Every species contains a unique and complex mixture of bioactive metabolites-phytoecdysteroids (PEs), iridoid glycosides, withanolides, neo-clerodane terpenoids, flavonoids, phenolics, and other chemicals with high therapeutic potential. Phytoecdysteroids, the main compounds of interest, are natural anabolic and adaptogenic agents that are widely used as components of dietary supplements. Wild plants remain the main source of Ajuga bioactive metabolites, particularly PEs, which leads to frequent overexploitation of their natural resources. Cell culture biotechnologies offer a sustainable approach to the production of vegetative biomass and individual phytochemicals specific for Ajuga genus. Cell cultures developed from eight Ajuga taxa were capable of producing PEs, a variety of phenolics and flavonoids, anthocyanins, volatile compounds, phenyletanoid glycosides, iridoids, and fatty acids, and demonstrated antioxidant, antimicrobial, and anti-inflammatory activities. The most abundant PEs in the cell cultures was 20-hydroxyecdysone, followed by turkesterone and cyasterone. The PE content in the cell cultures was comparable or higher than in wild or greenhouse plants, in vitro-grown shoots, and root cultures. Elicitation with methyl jasmonate (50-125 µM) or mevalonate and induced mutagenesis were the most effective strategies that stimulated cell culture biosynthetic capacity. This review summarizes the current progress in cell culture application for the production of pharmacologically important Ajuga metabolites, discusses various approaches to improve the compound yield, and highlights the potential directions for future interventions.

Flavonoids and Phenolic Acids from Aerial Part of Ajuga integrifolia (Buch.-Ham. Ex D. Don): Anti-Shigellosis Activity and In Silico Molecular Docking Studies.

Shigellosis is one of the major causes of death in children worldwide. Flavonoids and phenolic acids are expected to demonstrate anti-shigellosis activity and anti-diarrheal properties. The aerial part of A. integrifolia is commonly used against diarrhea. This study aimed to identify flavonoids and phenolic acids responsible for this therapeutic purpose. Antioxidant activity, total phenol content, and total flavonoid content were determined. The antibacterial activity of the aerial part against Shigella spp. was also tested using the agar well diffusion method. HPLC analysis was performed using UHPLC-DAD for different extracts of the aerial part. Autodock Vina in the PyRx platform was used to screen responsible components. Ciprofloxacin was used as a reference drug. An enzyme taking part in pyrimidine biosynthesis was used as a target protein. Molecular docking results were visualized using Discovery Studio and LigPlot1.4.5 software. Antioxidant activity, total phenol content, and total flavonoid content are more significant for the aerial part of A. integrifolia. From HPLC analysis, the presence of the flavonoids, quercetin, myricetin, and rutin and the phenolic acids gallic acid, chlorogenic acid, and syringic acid were identified from the aerial part of A. integrifolia. Regarding the antibacterial activity, the aerial part shows considerable activity against Shigella spp. Binding energies, RMSD and Ki values, interaction type, and distance are considered to identify the components most likely responsible for the therapeutic effects and observed activity. Antioxidant activity, total phenol content, and total flavonoid content of the aerial part are in line with anti-shigellosis activity. The top five components that are most likely potentially responsible for therapeutic purposes and anti-shigellosis activity are chlorogenic acid, rutin, dihydroquercetin, dihydromyricetin, and kaempferol.

Chemical constituents of Ajuga forrestii and their anti-ferroptosis activity.

Six new neoclerodane diterpenoids (1-6), along with ten known compounds (7-16), were isolated from Ajuga forrestii. Their structures were elucidated by HRESIMS, 1D and 2D NMR, ECD calculation, and single-crystal X-ray diffraction analysis. The structure of a known neoclerodane diterpene ajudecunoid C (6) was revised based on the reported NMR empirical rules. All the isolates were evaluated for their inhibitory effect on RSL3-induced ferroptosis in HT22 mouse hippocampal neuronal cells. Among them, compounds 8, 9, and 12 significantly inhibited RSL3-induced ferroptosis with EC50 values of 0.45 µM, 0.076 µM, and 0.14 µM.

New neo-Clerodane Diterpenoids Isolated from Ajuga decumbens Thunb., Planted at Pingtan Island of Fujian Province with the Potent Anticancer Activity.

AIMS: The aim of this study is to find the anticancer lead compounds or drug candidates from Chinese Traditional Plant Medicine of Ajuga decumbens Thunb. BACKGROUND: Ajuga decumbens Thunb. has been used in clinical for a long time in China and was selected in "Chinses Pharmacopoeia" (part I in 1977) for its wide spectrum biological activities: such as anticancer, antioxidant, antifeedant, antibacterial, anti-inflammatory, antihyperlipidemic, anti-cholinesterase and cytotoxicity activities. However, there are relatively fewer studies of Ajuga decumbens Thunb. that have been carried out till now. For some years, our research group focused on the discovery of new anticancer agents, so we studied the chemical compositions of Ajuga decumbens Thunb., planted in Pingtan island of Fujian Province, to discover new anticancer lead compounds or candidates from this Chinese Traditional Plant Medicine. METHODS: The dichloromethane (DCM) extract was obtained in this work, and then this extract was used for silica gel column chromatography to obtain different polar fractions. Several similar fractions were combined according to TLC or HPLC analysis. The combined fractions were isolated by preparative TLC or preparative HPLC to obtain the pure compounds and HPLC was used to detect the purity. All isolated compounds were determined by NMR (1HNMR, 13CNMR, DEPT, HMBC, HSQC, 1H-1H COSY and NOESY), HRESIMS and single crystal X-ray diffraction methods. The in vitro anticancer activity was evaluated using CCK8 method. RESULTS: Seven compounds [three new compounds 1-3; and four known compounds (Ajugacumbins A, Ajugacumbin B, Ajugamarin A2 and Ajugamarin A1)] were isolated from Ajuga decumbens Thunb. in this work, and their structures were confirmed. The biological evaluation showed that 3 and Ajugamarin A1 exhibited potent in vitro anticancer activity both against A549 cell lines with IC50s=71.4 µM and 76.7 µM; and against Hela cell lines with IC50s=71.6 mM and 5.39×10-7 µM, respectively. CONCLUSION: Compounds (3 and Ajugamarin A1) can be regarded as the lead compounds for the development of anticancer agents.

Chemical Composition, Antioxidant, and Antimicrobial Activities of Leaves of Ajuga Iva.

The main objective of this research was to study the biological characteristics in terms of antioxidant and antimicrobial activities of Ajuga iva and determine the best analytical and extraction methods applicable to this specie and studied compounds. A short screening of its nutritional value in terms of chemical composition is also included. A. iva leaves were analyzed for crude protein (CP), cell wall [neutral detergent fiber (NDF), acid detergent fiber (ADF), and acid detergent lignin (ADL)], minerals, fatty acids, essential oils, and phenolic compounds. Mature aerial parts of A. iva were randomly collected during the Spring season from Mograne-Zaghouan, Tunisia. Leaves of A. iva contained 13.4 ± 0.4% CP, 26.3 ± 0.35% NDF, 20.2 ± 0.42% ADF, and 5.13 ± 0.21% ADL. Mineral content (13.0 ± 0.45%) was mainly composed of potassium (4.5% g DM) and magnesium (4.25% DM). Leaves of A. iva had linolenic (26.29 ± 0.760%) and linoleic (37.66 ± 2.35%) acids as the main components of the acid profile. Thymol was found to be the most dominant (23.43%) essential oil, followed by 4-vinylguaiacol (14.27%) and linalool (13.66%). HPLC-PDA-ESI-MS/MS analysis pointed out the presence of phytoecdysteroids. Phenolic acids and flavonoids, such as glycosylated derivatives of naringenin, eriodyctiol, and apigenin, were detected in the methanol extract of A. iva leaves. Our results underline the importance of choosing proper extraction methods and solvents to extract and characterize the described compounds profile of A. iva leaves. Results also show A. iva leaves as a potential source of functional ingredients with beneficial health-promoting properties. Overall, leaves of A. iva have low biological activities (antioxidant and antimicrobial activities) with a chemical composition suitable as a feed for ruminants in rangeland pasture. It also has low-grade antibacterial or medicinal characteristics when fed to ruminants.

Polyphenol Rich Ajuga bracteosa Transgenic Regenerants Display Better Pharmacological Potential.

Ajuga bracteosa Wall. ex Benth. is an endangered medicinal herb traditionally used against different ailments. The present study aimed to create new insight into the fundamental mechanisms of genetic transformation and the biological activities of this plant. We transformed the A. bracteosa plant with rol genes of Agrobacterium rhizogenes and raised the regenerants from the hairy roots. These transgenic regenerants were screened for in vitro antioxidant activities, a range of in vivo assays, elemental analysis, polyphenol content, and different phytochemicals found through HPLC. Among 18 polyphenolic standards, kaempferol was most abundant in all transgenic lines. Furthermore, transgenic line 3 (ABRL3) showed maximum phenolics and flavonoids content among all tested plant extracts. ABRL3 also demonstrated the highest total antioxidant capacity (8.16 ± 1 µg AAE/mg), total reducing power, (6.60 ± 1.17 µg AAE/mg), DPPH activity (IC50 = 59.5 ± 0.8 µg/mL), hydroxyl ion scavenging (IC50 = 122.5 ± 0.90 µg/mL), and iron-chelating power (IC50 = 154.8 ± 2 µg/mL). Moreover, transformed plant extracts produced significant analgesic, anti-inflammatory, anticoagulant, and antidepressant activities in BALB/c mice models. In conclusion, transgenic regenerants of A. bracteosa pose better antioxidant and pharmacological properties under the effect of rol genes as compared to wild-type plants.

Combined Effects of Exercise and Phytoanabolic Extracts in Castrated Male and Female Mice.

Dry extracts from the Eurasian plants, Ajuga turkestanica, Eurycoma longifolia, and Urtica dioica have been used as anabolic supplements, despite the limited scientific data on these effects. To assess their actions on early sarcopenia signs, male and female castrated mice were supplemented with lyophilized extracts of the three plants, isolated or in association (named TLU), and submitted to resistance exercise. Ovariectomy (OVX) led to body weight increase and non-high-density cholesterol (HDL) cholesterol elevation, which had been restored by exercise plus U. dioica extract, or by exercise and TLU, respectively. Orchiectomy (ORX) caused skeletal muscle weight loss, accompanied by increased adiposity, being the latter parameter reduced by exercise plus E. longifolia or U. dioica extracts. General physical activity was improved by exercise plus herbal extracts in either OVX or ORX animals. Exercise combined with TLU improved resistance to fatigue in OVX animals, though A. turkestanica enhanced the grip strength in ORX mice. E. longifolia or TLU also reduced the ladder climbing time in ORX mice. Resistance exercise plus herbal extracts partly altered gastrocnemius fiber size frequencies in OVX or ORX mice. We provide novel data that tested ergogenic extracts, when combined with resistance exercise, improved early sarcopenia alterations in castrated male and female mice.

Ajuga iva water extract antihypertensive effect on stroke-prone spontaneously hypertensive rats, vasorelaxant effects ex vivo and in vitro activity of fractions.

ETHNOPHARMACOLOGICAL RELEVANCE: Ajuga iva (L.) Schreb. (Labiatae) (AI) is used in folk medicine for a variety of ailments, including diabetes mellitus and hypertension. AIM OF THE STUDY: In this work, we aimed to investigate the antihypertensive and vasorelaxant effects of AI aqueous extract in stroke prone spontaneously hypertensive rats (SHR-SP). MATERIAL AND METHODS: Male SHR-SP rats were orally force-fed AI aqueous extract (500 mg/kg body weight) daily for one week. Systolic blood pressure and urine output were recorded in vivo by non-invasive methods. AI vasoactive effects on noradrenaline contractile response and acetylcholine-evoked relaxation were assessed ex vivo on aorta rings of treated and untreated SHR-SP rats. AI extract was then subjected to bio-guided fractionation using solvents of increasing polarity. For each fraction, in vitro vasorelaxation assay was performed on noradrenaline-precontracted aorta of Wistar rats, in the absence/presence of N-nitro-L-arginine (L-NNA). HPLC analysis of AI total extract, and the most in vitro active AI residual aqueous extract fraction (A1) was performed using naringin, naringenin, apigenin, apigenin 7-O-glucoside as marker compounds. RESULTS: AI aqueous extract (500 mg/kg) significantly (P < 0.05) decreased systolic blood pressure (SBP) in SHR-SP rats, while not affecting the urine output. In ex vivo experiments, the total extract decreased contractile response to noradrenaline of aortic rings isolated from AI-treated SHR-SP rats with or without addition of N-nitro-L-arginine, but endothelium dependent relaxation evoked by acetylcholine in noradrenaline-contracted aortic rings was not affected by the extract treatment. In vitro experiments on AI aqueous extract fractions showed that its polar fraction was the only one affecting in vitro noradrenaline induced contractions, but only in an endothelium dependent manner. This fraction was shown by HPLC-UV to contain flavonoid glycosides among other polar compounds whose activity and mode of action may be modified in vivo by metabolization. CONCLUSION: These results support the use of AI as antihypertensive treatment in folk medicine. The systolic blood pressure decrease may be attributed at least in part to vasorelaxant glycosylated/polar phenolic compounds as flavonoids and/or their metabolites.

Diterpenoids from the Whole Plants of Ajuga nipponensis and Their Inhibition of RANKL-Induced Osteoclastogenesis.

Two new diterpenoids, ajudecunoid A (1) and ajudecunoid B (14), along with thirteen known diterpenoids, were isolated from the whole plants of Ajuga nipponensis Makino. Their structures were elucidated by the extensive spectroscopic analysis (UV, IR, MS, and NMR). The absolute configurations of ajudecunoid A (1) and ajudecunoid B (14) were defined through analysis of X-ray crystallography. Fifteen compounds were evaluated for inhibition of the formation of osteoclasts in bone marrow-derived macrophages (BMM) cells. Two neo-clerodane diterpenoids ajuganipponin B (5) and (12S)-6α,19-diacetoxy-18-chloro-4α-hydroxy-12-tigloyloxy-neo-clerod-13-en-15,16-olide (12) showed significant inhibition of osteoclastogenesis with IC50 values of 0.88 and 0.79 µM, respectively. Here we firstly reported diterpenoids with anti-osteoclastogenesis activity from A. nipponensis.

Beneficial effects of Ajuga chamaepitys (L.) Schreber subsp. chia (Schreber) and its iridoids on the colitis model: Histopathological and biochemical evidence.

In Turkish folk medicine, aerial parts of Ajuga chamaepitys (L.) Schreber subsp. chia (Schreber) are used for the treatment of diarrhea. The crude methanolic extract of aerial parts of A. chamaepitys subsp. chia was sequentially fractionated into five subextracts; n-hexane, dichloromethane, ethyl acetate, n-butanol and aqueous extracts. Effects of the methanol extract, subextracts and fractions were investigated in acetic acid-induced rat colitis model. The MeOH extract and n-BuOH subextract have regulated the caspase-3, myeloperoxidase, TNF-α, IL-6 levels and antioxidant parameters. After confirmation of the activity against ulcerative colitis, n-BuOH subextract was subjected to more chromatographic separation for the isolation of compounds ajugoside (1), asperulosidic acid (2) and deacetyl-asperulosidic acid (3). As a conclusion, A. chamaepitys subsp. chia can be used in cell, tissue, or individual-specific treatments that will be developed in the future treatment of IBD, or as a complementary therapeutic agent that contributes to these treatments.

Ethnomedicinal use, phytochemistry, pharmacology, and toxicology of Ajuga iva (L.,) schreb.

ETHNOPHARMACOLOGICAL RELEVANCE: Ajuga iva (L.,) Schreb (A. iva). is a medicinal plant commonly used in Africa to treat several diseases such as diabetes, rheumatism, allergy, cancer, renal, metabolic disorders, cardiovascular disorders, digestive, and respiratory disorders. AIM OF THE REVIEW: We highlighted previous reports on A. iva including its ethnopharmacological uses, the chemistry of its secondary metabolites, in vitro and in vivo pharmacological properties, and toxicological evidence. MATERIALS AND METHODS: The data on A. iva were gathered using scientific research databases such as ScienceDirect, PubMed, SpringerLink, Web of Science, Scopus Wiley Online, and Google Scholar. In this review, studies focused on A. iva and its phytopharmacological activities were explored. RESULTS: A. iva is used by many North African folk medicine practitioners especially against diabetes and immunological diseases. Our analysis of the previous reports confirmed the scientific evidence of A. iva ethnomedicinal uses, especially the antidiabetic and anti-hypercholesterolemia activity. However, there was no clear correlation between previous pharmacological reports on A. iva and its other ethnomedicinal uses in the treatment of rheumatism, allergy, metabolic, digestive, and respiratory disorders. The extracts and isolated compounds from A. iva exhibited numerous in vitro and in vivo pharmacological activities such as antidiabetic, antioxidant, antimicrobial, anti-hypercholesterolemia, insecticide, and litholitic effects. Chemical characterization using GC-MS, HPLC, and NMR revealed the presence of many chemical compounds such as 20-hydroxyecdysone, cyasterone, ajugasterone, apigenin dihexoside, apigenin, carvacrol, ecdysterone, palmitic acid in different parts of A. iva. These compounds belong to different classes of chemical compounds such as steroids, flavonoids, fatty acids, and terpenoids. CONCLUSIONS: A. iva extracts especially from the leaves showed significant antidiabetic, antioxidant, anti-hypercholesterolemia, and analgesic effects. Future studies are required to validate the results of clinical trials on A. iva antidiabetic, anti-hypercholesterolemia, antioxidant/anti-inflammatory, antimicrobial, and analgesic properties. Toxicological validation and pharmacokinetics investigation are necessary to validate the efficacy and safety A. iva extracts and its secondary metabolites. An in-depth investigation is needed to reveal the biological activity of A. iva active compounds in preventing the development of cancer and neurodegenerative disorders such as Alzheimer's and Parkinson's diseases.

Anti-inflammatory neo-Clerodane Diterpenoids from Ajuga pantantha.

Eight new neo-clerodane diterpenoids (1-8) were acquired from the aerial parts of Ajuga pantantha. Spectroscopic data analysis permitted the definition of their structures, and experimental and calculated electronic circular dichroism data were used to define their absolute configurations. Compounds 2 and 4-8 were found to have NO inhibitory effects with IC50 values of 20.2, 45.5, 34.0, 27.0, 45.0, and 25.8 µM, respectively. The more potent compounds 2, 6, and 8 were analyzed to establish their anti-inflammatory mechanism, including regulation of the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) proteins as well as their binding interactions with the two proteins.

Synergistic Antibacterial Efficacy of Biogenic Synthesized Silver Nanoparticles using Ajuga bractosa with Standard Antibiotics: A Study Against Bacterial Pathogens.

BACKGROUND: Multi-drug resistance in bacterial pathogens is a major concern of today. Green synthesis technology is being used to cure infectious diseases. OBJECTIVES: The aim of the current research was to analyze the antibacterial, antioxidant, and phytochemical screening of green synthesized silver nanoparticles using Ajuga bracteosa. METHODS: Extract of A. bracteosa was prepared by maceration technique. Silver nanoparticles were synthesized using A. bracteosa extract and were confirmed by UV-Vis spectrophotometer, Scanning Electron Microscope (SEM) and Fourier Transform Infrared Spectroscopy (FTIR). The antibacterial, anti-biofilm, cell proliferation inhibition, TLC-Bioautography, TLC-Spot screening, antioxidant, and phytochemical screening were also investigated. RESULTS: UV-Vis spectrum and Scanning electron microscopy confirmed the synthesis of green nanoparticles at 400 nm with tube-like structures. FTIR spectrum showed that functional groups of nanoparticles have a role in capping and stability of AgNP. Agar well diffusion assay represented the maximum antibacterial effect of ABAgNPs against Escherichia coli, Klebsiella pneumoniae, Streptococcus pyogenes, Staphylococcus aureus, and Pseudomonas aeruginosa at 0.10 g/mL concentration compared to ABaqu. Two types of interactions among nanoparticles, aqueous extract, and antibiotics (Synergistic and additive) were recorded against tested pathogens. Crystal violet, MTT, TLC-bio-autography, and spot screening supported the findings of the antibacterial assay. Highest antioxidant potential effect in ABaqu was 14.62% (DPPH) and 13.64% (ABTS) while 4.85% (DPPH) and 4.86% (ABTS) was recorded in ABAgNPs. Presence of phytochemical constituents showed pharmacological importance. CONCLUSION: It was concluded that green synthesis is an innovative technology in which natural products are conjugated with metallic particles and are used against infectious pathogens. The current research showed the significant use of green nanoparticles against etiological agents.

A novel Ursane-type triterpene saponin from Ajuga multiflora.

A phytochemical study was carried out on Ajuga multiflora, and obtained a novel ursane-type triterpene saponin, i.e., 3-epi-ursolic acid 3-O-palmitoyl 28-O-α-L-arabinofuranosyl ester (1), named ajugoside. Its structure and relative configuration were identified mainly by spectroscopic methods including 1D- and 2D-NMR techniques as well as HR-MS analysis. This paper described the isolation and structural elucidation of ajugoside (1) for the first time.

Ethnomedicinal Uses, Phytochemistry, Pharmacology, and Toxicology of Species from the Genus Ajuga L.: A Systematic Review.

The present review is aimed at providing a comprehensive summary of the botanical characteristics, ethnomedicinal uses, phytochemical, pharmacological, and toxicological studies of the genus Ajuga L. The extensive literature survey revealed Ajuga L. species to be a group of important medicinal plants used for the ethnomedical treatment of rheumatism, fever, gout, sclerosis, analgesia, inflammation, hypertension, hyperglycemia, joint pain, palsy, amenorrhea, etc., although only a few reports address the clinical use and toxicity of these plants. Currently, more than 280 chemical constituents have been isolated and characterized from these plants. Among these constituents, neo-clerodane diterpenes and diterpenoids, phytoecdysteroids, flavonoids, and iridoids are the major bioactive compounds, possessing wide-reaching biological activities both in vivo and in vitro, including anti-inflammatory, antinociceptive, antitumor, anti-oxidant, antidiabetic, antimicrobial, antifeedant, antidiarrhoeal, hypolipidemic, diuretic, hypoglycaemic, immunomodulatory, vasorelaxant, larvicidal, antimutagenic, and neuroprotective activity. This review is aimed at summarizing the current knowledge of the ethnomedicinal uses, phytochemistry, biological activities, and toxicities of the genus Ajuga L. to reveal its therapeutic potentials, offering opportunities for future researches. Therefore, more focus should be paid to gathering information about their toxicology data, quality-control measures, and the clinical application of the bioactive ingredients from Ajuga L. species.

Comparative Phytochemical Profile, Antioxidant, Antimicrobial and In Vivo Anti-Inflammatory Activity of Different Extracts of Traditionally Used Romanian Ajuga genevensis L. and A. reptans L. (Lamiaceae).

Several Ajuga species are used in Romanian folk medicine for their antioxidant, antimicrobial and anti-inflammatory properties, to treat pain, fever or arthritis. Still, the active compounds responsible for these effects and their mechanism of action are scarcely known. This research was designed to investigate the phytochemical profile (e.g. iridoids, polyphenolic compounds, phytosterols), as well as the biological potential (antioxidant, antibacterial, antifungal, anti-inflammatory properties) of two selected Ajuga species collected from different regions of Romanian spontaneous flora. The main compounds identified in A. reptans aerial parts extracts were 8-O-acetylharpagide, isoquercitrin and ß-sitosterol, whilst in A. genevensis were 8-O-acetylharpagide, luteolin and campesterol. The extracts were screened for their antioxidant potential using different methods (DPPH, TEAC, EPR) and the results showed a good activity, in accordance with the polyphenol content (18-26 mg GAE/g dw). The antifungal activity on the tested strains was good. The determination of few parameters linked with the inflammatory mechanism allowed the assessment of in vivo anti-inflammatory potential. Ajuga reptans and A. genevensis ethanol extracts had anti-inflammatory activity through lowering the oxidative stress, phagocytosis, PMN and total leukocytes. The best anti-oxidative and anti-inflammatory activity was observed for the Ajuga reptans 100 mg dw/mL extract when compared with diclofenac, thus the dose could be correlated with the pharmacological effect. These findings provide substantial evidence that both selected Ajuga species have the potential to be valued as sources of phytochemicals in effective anti-inflammatory herbal preparations.

A new abietane diterpenoid glycoside from ajuga ovalifolia var. calantha.

A new abietane diterpenoid glycoside, ajugaside B (1), along with three known compounds (2-4), were isolated from the whole plants of Ajuga ovalifolia var. calantha. The structure of the new compound (1) was elucidated by means of spectroscopic analyses (HRESIMS, IR, NMR and ECD). All of the isolated compounds were evaluated for their antitumor activities against MGC803, MCF-7, A549, HT29 and HepG2 cell lines. Compounds 3-4 showed moderate cytotoxicity against all tested cell lines with IC50 values of 1.8-7.3 µM.

Development of a NIR Method for the In-Line Quantification of the Total Polyphenolic Content: A Study Applied on Ajuga genevensis L. Dry Extract Obtained in a Fluid Bed Process.

This study describes an innovative in-line near-infrared (NIR) process monitoring method for the quantification of the total polyphenolic content (TPC) of Ajuga genevensis dry extracts. The dry extract was obtained in a fluidized bed processor, by spraying and adsorbing a liquid extract onto an inert powder support. NIR spectra were recorded continuously during the extract's spraying process. For the calibration of the in-line TPC quantification method, samples were collected during the entire process. The TPC of each sample was assessed spectroscopically, by applying a UV-Vis reference method. The obtained values were further used in order to develop a quality OPLS prediction model by correlating them with the corresponding NIR spectra. The final dry extract registered good flowability and compressibility properties, a concentration in active principles three times higher than the one of the liquid extract and an overall process yield of 85%. The average TPC's recovery of the NIR in-line prediction method, compared with the reference UV-Vis one, was 98.7%, indicating a reliable monitoring method which provided accurate predictions of the TPC during the process, permitting a good process overview and enabling us to establish the process's end point at the exact moment when the product reaches the desired TPC concentration.