Analyzing the Chemical Constituents and In Vitro Biological Effects of Extracts Artemisia absinthium L. Extracts Obtained Using Different Methodologies and Solvents.

The present study aimed to investigate the chemical constituents of different extracts from aerial parts of A. absinthium and to evaluate their antioxidant and enzyme inhibition activity. Extracts were prepared by maceration, infusion or Soxhlet techniques. Results showed that the highest total phenolic and flavonoids contents was recorded respectively from the hexane extract prepared by maceration and ethyl acetate extract obtained by Soxhlet method. The characteristic compounds of Artemisia species artemetin, casticin, sesartemin and yangambin in addition to coumarins were identified in all extracts. Aqueous extract obtained by infusion exerted the highest radical scavenging and ions reducing properties while that prepared by maceration displayed the highest chelating power. Methanol extracts obtained by the two methods of extraction exerted the highest anti-Tyr activity while that obtained by maceration showed the best α-glucosidase inhibition activity. These findings indicated that A. absinthium is a rich source of bioactive molecules with possible therapeutic applications.

Exploring anti-inflammatory and antioxidant-related quality markers of Artemisia absinthium L. based on spectrum-effect relationship.

INTRODUCTION: Artemisia absinthium L. is a well-known medicinal, aromatic, and edible plant with important medicinal and economic properties and a long history of use in treating liver inflammation and other diseases; however, there has been insufficient progress in quality control. OBJECTIVE: This study aimed to investigate the quality markers for the anti-inflammatory and antioxidant activities of A. absinthium based on spectrum-effect relationship analysis. MATERIALS AND METHODS: Eighteen batches of A. absinthium from different origins were used. Chemical fingerprints were obtained by ultra-performance liquid chromatography (UPLC). The chemical compositions were identified by quadrupole-Orbitrap high-resolution mass spectrometry. Anti-inflammatory activity was assessed by inhibition of cyclooxygenase-2 and 15-lipoxygenase in vitro and inhibition of nitric oxide release in lipopolysaccharide-induced BV-2 cells. Antioxidant activity was assessed by DPPH and ABTS radical scavenging assays. The relationship between bioactivity and chemical fingerprints was then analyzed using chemometrics including gray relational analysis, bivariate correlation analysis, and orthogonal partial least squares analysis. RESULTS: Different batches of A. absinthium extracts possessed significant anti-inflammatory and antioxidant activities to varying degrees. Eighty compounds were identified from A. absinthium, and 12 main common peaks were obtained from the UPLC fingerprints. P3 (chlorogenic acid), P5 (isochlorogenic acid A), and P6 (isochlorogenic acid C) were screened as the most promising active compounds by correlation analysis and further validated for their remarkable anti-inflammatory effects. CONCLUSION: This is the first study to screen the quality markers of A. absinthium by establishing the spectrum-effect relationship, which can provide a reference for the development of quality standards and further research on A. absinthium.

Total Mineral Content in Raw Materials of Ten Medicinal Plants from Roadside Biotopes of Voronezh Oblast.

The total mineral content was studied in medicinal plants from roadside and railside biotopes of the Voronezh region. Pharmacopoeial plant raw materials of 10 species were evaluated: roots of Taraxacum officinale F.H. Wigg and Arctium lappa L.; herb of Polygonum aviculare L., Artemisia absinthium L., Leonurus quinquelobatus Gilib., and Achillea millefolium L.; leaves of Urtica dioica L. and Plantago major L.; and flowers of Tanacetum vulgare L. and Tilia cordata Mill. Plant raw materials were collected near roads and railways of various types in the periods specified in regulatory documents. The total ash content in plant material was used to determine the minimum allowable distances from various roads and railways for collecting plant material. The minimum allowable distance from heavy-traffic motorways was recommended to be 210 m in the forest zone, 240 m in the forest-steppe zone, and 380 m in the steppe zone. A distance of at least 80 m was recommended for secondary low-speed roads and railways.

Removal of Pb ions using green Co3O4 nanoparticles: Simulation, modeling, adsorption, and biological studies.

Chemical co-precipitation synthesized novel and green cobalt-oxide nanoparticles (Co3O4-NPs) utilizing cobalt nitrate as cobalt precursors. FTIR, Raman, scanning electron microscopy, UV visible, X-ray powder diffraction, and BET was used to analyze the surface characteristics, composition, and morphology, of the NPs. These green Co3O4-NPs were employed to remove Pb ions from simulated wastewater solutions at various pH, adsorbate, temperature, and dose concentrations. At dose 20 mg/L, pH 6.0, 20 mg/L (Pb(II) solution, 25 °C of temperature, and 45 min for equilibrium, nearly 99.44% of Pb ions were removed. To evaluate the kinetic data, four different kinetic equations were used. The data fit the Elovich rate equation better than the other three models. Thermodynamic and isothermal studies were also evaluated, and the maximum adsorption capacity of 450.45 mg/g was observed at 298.15 K. 0.1 M HNO3, and 0.1 HCl were used to regenerate used Co3O4-NPs. Simulation results show the strong correlation of the Co atom in the Co3O4-NPs generates active delocalized surface states, which are energetically most favorable for heavy metal (Pb ions) adsorption and removal, supporting the experimental outcomes. In concluding remarks, green Co3O4-NPs can also be used as an adsorbent to remove Pb ions from wastewater bodies.

New Evidence for Artemisia absinthium as an Alternative to Classical Antibiotics: Chemical Analysis of Phenolic Compounds, Screening for Antimicrobial Activity.

Artemisia absinthium, an important herb of the Artemisia genus, was evaluated in this study for its potential as an alternative to classical antibiotics. The antimicrobial activity of methanol extracts of A. absinthium (MEAA) was evaluated using the broth microdilution method, revealing that A. absinthium exhibited broad-spectrum antibacterial and antifungal activity. Ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry (UPLC-Q-TOF-MS) was used to analyze the chemical profile of the MEAA, with a focus on flavonoids, quinic acids, and glucaric acids. A total of 90 compounds were identified, 69 of which were described for the first time in A. absinthium. Additionally, a new class of caffeoyl methyl glucaric acids was identified. The main active compounds were quantified and screened for antimicrobial activity. A. absinthium was found to be rich in quinic acids and flavonoids. The screening for antimicrobial activity also revealed that salicylic acid, caffeic acid, casticin, and 3,4-dicaffeoylquinic acid had varying degrees of antimicrobial activity. The acute toxicity of MEAA was examined following OECD guidelines. The administration of 5000 mg/kg bw of MEAA did not result in mortality in male and female mice. Furthermore, there were no observed effects on the visceral organs or general behavior of the mice, demonstrating the good safety of MEAA. This study provides new evidence for the use of A. absinthium as an alternative to classical antibiotics in addressing the problem of bacterial resistance.

Total Flavonoids in Artemisia absinthium L. and Evaluation of Its Anticancer Activity.

To overcome the shortcomings of traditional extraction methods, such as long extraction time and low efficiency, and considering the low content and high complexity of total flavonoids in Artemisia absinthium L., in this experiment, we adopted ultrasound-assisted enzymatic hydrolysis to improve the yield of total flavonoids, and combined this with molecular docking and network pharmacology to predict its core constituent targets, so as to evaluate its antitumor activity. The content of total flavonoids in Artemisia absinthium L. reached 3.80 ± 0.13%, and the main components included Astragalin, Cynaroside, Ononin, Rutin, Kaempferol-3-O-rutinoside, Diosmetin, Isorhamnetin, and Luteolin. Cynaroside and Astragalin exert their cervical cancer inhibitory functions by regulating several signaling proteins (e.g., EGFR, STAT3, CCND1, IGFIR, ESR1). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis showed that the anticancer activity of both compounds was associated with the ErbB signaling pathway and FoxO signaling pathway. MTT results showed that total flavonoids of Artemisia absinthium L. and its active components (Cynaroside and Astragalin) significantly inhibited the growth of HeLa cells in a concentration-dependent manner with IC50 of 396.0 ± 54.2 µg/mL and 449.0 ± 54.8 µg/mL, respectively. Furthermore, its active components can mediate apoptosis by inducing the accumulation of ROS.

In Vitro Assessment of the Antidiabetic and Anti-Inflammatory Potential of Artemisia absinthium, Artemisia vulgaris and Trigonella foenum-graecum Extracts Processed Using Membrane Technologies.

Recently, there has been increased interest in the discovery of new natural herbal remedies for treating diabetes and inflammatory diseases. In this context, this work analyzed the antidiabetic and anti-inflammatory potential of Artemisia absinthium, Artemisia vulgaris and Trigonella foenum-graecum herbs, which have been studied less from this point of view. Therefore, extracts were prepared and processed using membrane technologies, micro- and ultrafiltration, to concentrate the biologically active principles. The polyphenol and flavone contents in the extracts were analyzed. The qualitative analysis of the polyphenolic compounds was performed via HPLC, identifying chlorogenic acid, rosmarinic acid and rutin in A. absinthium; chlorogenic acid, luteolin and rutin in A. vulgaris; and genistin in T. foenum-graecum. The antidiabetic activity of the extracts was analyzed by testing their ability to inhibit α-amylase and α-glucosidase, and the anti-inflammatory activity was analyzed by testing their ability to inhibit hyaluronidase and lipoxygenase. Thus, the concentrated extracts of T. foenum-graecum showed high inhibitory activity on a-amylase-IC50 = 3.22 ± 0.3 µg/mL-(compared with acarbose-IC50 = 3.5 ± 0.18 µg/mL) and high inhibitory activity on LOX-IC50 = 19.69 ± 0.52 µg/mL (compared with all standards used). The concentrated extract of A. vulgaris showed increased α-amylase inhibition activity-IC50 = 8.57 ± 2.31 µg/mL-compared to acarbose IC50 = 3.5 ± 0.18 µg/mL. The concentrated extract of A. absinthium showed pronounced LOX inhibition activity-IC50 = 19.71 ± 0.79 µg/mL-compared to ibuprofen-IC50 = 20.19 ± 1.25 µg/mL.

Artemisia Species with High Biological Values as a Potential Source of Medicinal and Cosmetic Raw Materials.

Artemisia species play a vital role in traditional and contemporary medicine. Among them, Artemisia abrotanum, Artemisia absinthium, Artemisia annua, Artemisia dracunculus, and Artemisia vulgaris are the most popular. The chemical composition and bioactivity of these species have been extensively studied. Studies on these species have confirmed their traditional applications and documented new pharmacological directions and their valuable and potential applications in cosmetology. Artemisia ssp. primarily contain sesquiterpenoid lactones, coumarins, flavonoids, and phenolic acids. Essential oils obtained from these species are of great biological importance. Extracts from Artemisia ssp. have been scientifically proven to exhibit, among others, hepatoprotective, neuroprotective, antidepressant, cytotoxic, and digestion-stimulating activities. In addition, their application in cosmetic products is currently the subject of several studies. Essential oils or extracts from different parts of Artemisia ssp. have been characterized by antibacterial, antifungal, and antioxidant activities. Products with Artemisia extracts, essential oils, or individual compounds can be used on skin, hair, and nails. Artemisia products are also used as ingredients in skincare cosmetics, such as creams, shampoos, essences, serums, masks, lotions, and tonics. This review focuses especially on elucidating the importance of the most popular/important species of the Artemisia genus in the cosmetic industry.

Artemisia absinthium improves spatial performance and neuronal injury induced by amyloid- beta in the CA1 hippocampal area of male Wistar rats.

Alzheimer's disease is a neurodegenerative disorder. It is characterized by the presence of two aberrant structures in the brain, those are, amyloid plaques and neurofibrillary tangles, along with neuronal death. Amyloid-beta further exacerbates the metabolic decline and results in cognitive impairments. Because of the favorable antioxidant and anti-inflammatory activities of Artemisia absinthium (wormwood), this study aimed to evaluate the effects of hydroalcoholic extract of this plant on spatial memory performance, neuronal injury, and apoptosis induced by amyloid-beta. Forty-eight male Wistar rats (220-250 g) were divided into the following groups: 1) control; 2) sham (solvent; ICV); 3) amyloid-beta 1-40 (ICV); and 4) amyloid-beta plus A. absinthium (10, 50, and 100 mg/kg/day; gavage). Congo red and TUNEL staining were performed to investigate the neuronal injury. Also, the Morris water maze (MWM) test was used to evaluate the spatial memory of the experimental groups. The results showed that spatial memory for finding the hidden platform in the MWM task decreased significantly in the amyloid-beta group, compared to the control and sham groups. In contrast, treatment with A. absinthium improved spatial memory dose-dependently and reduced tissue degeneration, amyloid plaques, and apoptosis. It seems that the hydroalcoholic extract of A. absinthium, due to its antioxidant and anti-inflammatory activities, can effectively reverse spatial memory deficits and reduce amyloid-beta plaques.

Essential oils from Artemisia species inhibit biofilm formation and the virulence of Escherichia coli EPEC 2348/69.

Enteropathogenic Escherichia coli E2346/69 (EPEC) has caused foodborne outbreaks worldwide and the bacterium forms antimicrobial-tolerant biofilms. The anti-biofilm formation of various components of essential oils extracted from selected medicinal plants were investigated and tested on EPEC and wild strains of E. coli. Oils extracted from the family Asteraceae and their major common constituents at 0.031 and 0.062% (V/v) were found to significantly inhibit biofilm formation without affecting the growth of planktonic cells. In addition, three plants belonging to this family (Artemisia herba alba, Artemisia campestris and Artemisia absinthium) played important roles in the antimicrobial activity. Interestingly, their essential oils reduced the ability of E. coli (the EPEC and K12 strains) to form a biofilm. The crystal violet reduction assay showed that the plant extracts tested reduced biofilm formation with the inhibition of bacterial attachment up to 45% for EPEC and 70% for E. coli K-12 after 24 h treatment at 0.62 mg ml-1, demonstrating that Artemisia oils had a high anti-biofilm activity on the bacteria tested. The results indicate that the locus of enterocyte effacement (LEE) acquired by horizontal transfer promotes the formation of the attaching and effacing (A/E) lesion and increases the capacity of the photogen strain (EPEC) to form a biofilm. The chemical composition of the volatile compounds was obtained by gas chromatography-mass spectrometry analysis, which showed that the essential oils consisted of thirty-four compounds. Chamazulene (39.21%), ß-pinene (32.07%), and α-thujone (29.39%) were the main constituents of the essential oils of A. herba alba, A. absinthium and A. campestris, respectively.

Evaluation of the phytotoxic effect of the essential oil from Artemisia absinthium.

The chemical profile and the phytotoxicity of Artemisia absinthium essential oil (EO) were investigated to evaluate its potential value as a biopesticide for food safety purposes. A total of 54 compounds were identified in A. absinthium EO, with the most abundant constituents being eucalyptol (25.59%), linalool (11.99%), and ß-myrcene (10.05%). The EO, linalool, and a mixture of three major components exhibited potent suppressive activity against four receiver species; however, eucalyptol and ß-myrcene showed a much weaker effect. Bioassay-guided fractionation led to the isolation of linalool as the major active compound responsible for the EO's phytotoxicity. Subsequent scanning electron microscopy (SEM) analysis revealed that linalool significantly inhibited root-hair formation and metaxylem development. This is the first report on the determination of linalool as the major active phytotoxic compound in A. absinthium EO, as well as the elucidation of its mechanism of phytotoxicity from the perspective of root structure changes in the receiver species. Our results suggest that both the EO and its major constituents have potential value as environmentally friendly herbicides.

Chloroplast genome sequences of Artemisia maritima and Artemisia absinthium: Comparative analyses, mutational hotspots in genus Artemisia and phylogeny in family Asteraceae.

Artemisia L. is a complex genus of medicinal importance. Publicly available chloroplast genomes of few Artemisia species are insufficient to resolve taxonomic discrepancies at species level. We report chloroplast genome sequences of two further Artemisia species: A. maritima (151,061 bp) and A. absinthium (151,193 bp). Both genomes possess typical quadripartite structure comprising of a large single copy, a small single copy and a pair of long inverted repeats. The two genomes exhibited high similarities in genome sizes, gene synteny, GC content, synonymous and non-synonymous substitutions, codon usage, amino acids frequencies, RNA editing sites, microsatellites, and oligonucleotide repeats. Transition to transversion ratio was <1. Maximum likelihood tree showed Artemisia a monophyletic genus, sister to genus Chrysanthemum. We also identified 20 highly polymorphic regions including rpoC2-rps2, trnR-UCU-trnG-UCC, rps18-rpl20, and trnL-UAG-rpl32 that could be used to develop authentic and cost-effective markers to resolve taxonomic discrepancies and infer phylogenetic relationships among Artemisia species.

Methanolic extract of Artemisia absinthium prompts apoptosis, enhancing expression of Bax/Bcl-2 ratio, cell cycle arrest, caspase-3 activation and mitochondrial membrane potential destruction in human colorectal cancer HCT-116 cells.

The Artemisia absinthium (AA), belongs to the Asteraceae family, is used as a therapeutic agent in traditional medicine in Iran. It is a rich source of biology-active compounds. However, the molecular mechanism of AA contributing to cell proliferation and apoptosis is still unknown. This study aims to assess the anticancer activity of the methanolic extract of A. absinthium (MEAA) against human colorectal cancer HCT-116 cell line. The cytotoxic effects of MEAA on HCT-116 cells was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay. The expression levels of BAX and BCL-2 in HCT-116 cell line were examined by qRT-PCR. Annexin V/PI-flow cytometry technique was used to detect the cell cycle and apoptosis. MMP was predicted by Rhodamine 123 staining, and caspase 3 activity was analyzed by ELISA. Western blot method was performed to detect the expression level of BAX, Bcl-2 and Caspase-3 proteins. The MTT test revealed MEAA reduced the viability of HCT-116 cells. The mRNA and protein levels of BAX increased, but those of BCL-2 decreased in MEAA-treated cells. MEAA also prompted cell cycle arrest and induced apoptosis. After adding MEAA, the protein level and activity of caspase 3 and MMP destruction significantly increased. MEAA predominantly prompted apoptosis in HCT-116 cells by activating the mitochondrial pathway.

Impact of Artemisia absinthium hydrolate extracts with nematicidal activity on non-target soil organisms of different trophic levels.

Natural pesticides are considered a good alternative to synthetic pesticides to reduce environmental impacts. However, biopesticides may have unknown effects on the environment, and can affect non-target organisms. In this study, the ecotoxicological effects of an aqueous extract (hydrolate) from Spanish populations of Artemisia absinthium (var. Candial) showing a promising biopesticide activity, were evaluated on non-target soil organisms from different trophic levels (natural microbial communities characterized through 16S rRNA gene sequencing, the earthworm Eisenia fetida and the plant Allium cepa). The hydrolate usually was considered as a by-product of the distillation to obtain essential oils. However, recently has been found to have nematicide properties. The hydrolate caused acute toxicity at values of LC50 of 3.87% v/v for A. cepa and 0.07 mL/g for E. fetida. All the concentrations except for the most diluted (1% v/v) reduced the bacterial physiological activity compared to controls (LC50 = 25.72% v/v after 24 h of exposure). The hydrolate also slightly altered the ability of the microbial community to degrade carbon substrates. These results indicate that the hydrolate from A. absinthium may affect the survival and metabolic abilities of key soil organisms.

The Extracts of Artemisia absinthium L. Suppress the Growth of Hepatocellular Carcinoma Cells through Induction of Apoptosis via Endoplasmic Reticulum Stress and Mitochondrial-Dependent Pathway.

Artemisia absinthium L. has pharmaceutical and medicinal effects such as antimicrobial, antiparasitic, hepatoprotective, and antioxidant activities. Here, we prepared A. absinthium ethanol extract (AAEE) and its subfractions including petroleum ether (AAEE-Pe) and ethyl acetate (AAEE-Ea) and investigated their antitumor effect on human hepatoma BEL-7404 cells and mouse hepatoma H22 cells. The cell viability of hepatoma cells was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The apoptosis, cell cycle, mitochondrial membrane potential (Δψm), and reactive oxygen species (ROS) were analyzed by flow cytometry. The levels of proteins in the cell cycle and apoptotic pathways were detected by Western blot. AAEE, AAEE-Pe, and AAEE-Ea exhibited potent cytotoxicity for both BEL-7404 cells and H22 cells through the induction of cell apoptosis and cell cycle arrest. Moreover, AAEE, AAEE-Pe, and AAEE-Ea significantly reduced Δψm, increased the release of cytochrome c, and promoted the cleavage of caspase-3, caspase-9, and poly(ADP-ribose) polymerase (PARP) in BEL-7404 and H22 cells. AAEE, AAEE-Pe, and AAEE-Ea significantly upregulated the levels of ROS and C/EBP-homologous protein (CHOP). Further, AAEE, AAEE-Pe, and AAEE-Ea significantly inhibited tumor growth in the H22 tumor mouse model and improved the survival of tumor mice without side effects. These results suggest that AAEE, AAEE-Pe, and AAEE-Ea inhibited the growth of hepatoma cells through induction of apoptosis, which might be mediated by the endoplasmic reticulum stress and mitochondrial-dependent pathway.

Romanian Wormwood (Artemisia absinthium L.): Physicochemical and Nutraceutical Screening.

Artemisia species are used worldwide for their antioxidant, antimicrobial and anti-inflammatory properties. This research was designed to investigate the phytochemical profile of two ethanolic extracts obtained from leaves and stems of A. absinthium L. as well as the biological potential (antioxidant activity, cytotoxic, anti-migratory and anti-inflammatory properties). Both plant materials showed quite similar thermogravimetric, FT-IR phenolic profile (high chlorogenic acid) with mild antioxidant capacity [ascorbic acid (0.02-0.1) > leaves (0.1-2.0) > stem (0.1-2.0)]. Alcoholic extracts from these plant materials showed a cytotoxic effect against A375 (melanoma) and MCF7 (breast adenocarcinoma) and affected less the non-malignant HaCaT cells (human keratinocytes) at 72 h post-stimulation and this same trend was observed in the anti-migratory (A375, MCF7 > HaCat) assay. Lastly, extracts ameliorated the pro-inflammatory effect of TPA (12-o-tetradecanoylphorbol-13-acetate) in mice ears, characterized by a diffuse neutrophil distribution with no exocytosis or micro-abscesses.

Evaluation of anticancer, antioxidant activity and phenolic compounds of Artemisia absinthium L. Extract.

In the treatment of cancer, which remains a fatal disease, increasingly successful treatment rates of alternative therapies using the power of plants have directed the scientific world towards natural plant resources. This study aimed to examine the anti-cancer and antioxidant properties and identify the phenolic content of the methanolic extract obtained from Artemisia absinthium L. (AR) species, which is used as folk-medicine. The antioxidant activity of the extract was investigated using total phenolics, flavonoids, ABTS and CUPRAC methods. Phenolic component analysis of the plant extract was performed by LC-MS/MS. The anti-cancer property of AR extract was investigated on human colon (DLD-1), endometrium (ECC-1) cancer cells and embryonic kidney (HEK-293) cells. Cytotoxic effects were defined with MTT, apoptotic activity with DNA fragmentation ELISA and AO/EB fluorescent staining, the genotoxic effect with the comet assay and the intracellular oxidative status with TAS and TOS methods. As a result of the study, it was determined that AR extract showed an antioxidant effect, and as a result of the content analysis made with LC-MS/MS, phenolic components were determined, the most abundant being chlorogenic acid, followed by quinic acid, cinnamic acid, rhoifolin and malic acide. AR extract showed cytotoxic activity on DLD-1 and ECC-1 cancer cells, while the cytotoxic effect on HEK-293 cells was determined to be low.  It was determined that by increasing the intracellular amount of free radicals on cancer cells, this led to DNA damage, which consequently led to apoptosis of the cancer cells.

Topical Effects of Artemisia Absinthium Ointment and Liniment in Comparison with Piroxicam Gel in Patients with Knee Joint Osteoarthritis: A Randomized Double-Blind Controlled Trial.

BACKGROUND: Pain alleviation and improvement of functional status are the main objectives in the treatment of osteoarthritis. Artemisia absinthium (AA) was used traditionally in reducing pain and inflammation. The aim of the present study was to compare the effects of topical formulations of AA and piroxicam gel (PG) among patients with knee osteoarthritis. METHODS: In total, 90 outpatients aged 30-70 years with the diagnosis of primary osteoarthritis in at least one knee were enrolled in a randomized double-blind clinical trial. The patients referred to the Rheumatology Clinic at Shahid Beheshti Hospital in Hamadan province during 2012-2013. The patients were randomly assigned into three groups, 30 patients per group, and respectively received AA ointment (AAO) 3%, AA liniment (AAL) 3%, and PG; three times daily (TID) for 4 weeks. The patients were visited at baseline, week 4, and week 6. The effectiveness criteria were pain severity which was assessed with a 10-point visual analog scale (VAS), the Western Ontario and McMaster Universities osteoarthritis index (WOMAC) for total pain score (WTPS), total physical function score (WTPFS), and total stiffness score (WTSS). Repeated measure ANOVA, paired t test and post hoc were used to compare variables. Statistical analysis was performed using the SPSS software, version 13.0 (SPSS Inc., Chicago, Illinois). RESULTS: All groups had similar patient demographics. The administration of PG significantly improved all tested criteria with no recurrence after discontinuing the treatment protocol. AAO alleviated all tested factors except for WTSS. Alleviation was comparable to PG. AAL only reduced pain factors (VAS, WTPS) in week 4 with recurrence in week 6. CONCLUSION: Administration of Artemisia ointment may have beneficial effects in the treatment of osteoarthritis. Trial Registration Number: IRCT201202123109N3.

Functional Analysis of Amorpha-4,11-Diene Synthase (ADS) Homologs from Non-Artemisinin-Producing Artemisia Species: The Discovery of Novel Koidzumiol and (+)-α-Bisabolol Synthases.

The production of artemisinin, the most effective antimalarial compound, is limited to Artemisia annua. Enzymes involved in artemisinin biosynthesis include amorpha-4,11-diene synthase (ADS), amorpha-4,11-diene 12-monooxygenase (CYP71AV1) and artemisinic aldehyde Δ(11)13 reductase (DBR2). Although artemisinin and its specific intermediates are not detected in other Artemisia species, we reported previously that CYP71AV1 and DBR2 homologs were expressed in some non-artemisinin-producing Artemisia plants. These homologous enzymes showed similar functions to their counterparts in A. annua and can convert fed intermediates into the following products along the artemisinin biosynthesis in planta These findings suggested a partial artemisinin-producing ability in those species. In this study, we examined genes highly homologous to ADS, the first committed gene in the pathway, in 13 Artemisia species. We detected ADS homologs in A. absinthium, A. kurramensis and A. maritima. We analyzed the enzymatic functions of all of the ADS homologs after obtaining their cDNA. We found that the ADS homolog from A. absinthium exhibited novel activity in the cyclization of farnesyl pyrophosphate (FPP) to koidzumiol, a rare natural sesquiterpenoid. Those from A. kurramensis and A. maritima showed similar, but novel, activities in the cyclization of FPP to (+)-α-bisabolol. The unique functions of the novel sesquiterpene synthases highly homologous to ADS found in this study could provide insight into the molecular basis of the exceptional artemisinin-producing ability in A. annua.

Prenatal heat stress reduces male anogenital distance at birth and adult testis size, which are rescued by concurrent maternal Artemisia absinthium consumption.

Boars from sows with elevated plasma cortisol during pregnancy have shorter anogenital distance (AGD), a trait associated with subfertility. Since gestating sows often experience summer heat stress (HS), a mouse model was used to evaluate the effect of prenatal HS on AGD and fertility; efficacy of the heat stress-mitigating supplement Artemisia absinthium (AB) was also evaluated. Dams were treated from d 8-18 of gestation, residing in ambient temperatures from 0700 to 1900h. From 1900 to 0700h females were exposed to 34.13±0.27°C (HS) with access to water (HSW; n=9), HS with access to a 1% w/v decoction of AB (HSA; n=9), 20.81±0.20°C (thermal neutral; TN) with water (TNW; n=10) or TN with AB (TNA; n=10). Daily liquid consumption was measured from d 6-18, and tail temperature was recorded at 0700 and 1900h from d 8-18. Progeny weight and AGD were recorded at birth and weaning. At maturity, males were mated to non-littermate females from each treatment; these females were euthanized after 16 d of TN gestation. Reproductive traits were compared among all male/female treatment combinations; testes were weighed. Average liquid intake differed among treatments with HS and AB animals drinking more (P<0.0004). A treatment by time interaction for tail temperature (P<0.001) was observed; HS increased tail temperature of HSA and HSW animals similarly compared to TNA and TNW. Treatment affected (P<0.01) male birth AGD (HSW shortest; P<0.07). At maturity, HSW males also had the smallest testes (P<0.02). However, we observed no differences in fertility (P>0.10). These data indicate that in utero HS decreases both male birth AGD and adult testis size, effects which maternal AB consumption mitigates.