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Valeriana officinalis (Valerian) root extracts have been used by European and Asian cultures for millennia for their anxiolytic and sedative properties. However, the efficacy of these extracts suffers from variable yields and composition, making these extracts a prime candidate for microbial production. Recently, valerenic acid, a C15 sesquiterpenoid, was identified as the active compound that modulates the GABAA channel. Although the first committed step, valerena-4,7(11)-diene synthase, has been identified and described, the complete valerenic acid biosynthetic pathway remains to be elucidated. Sequence homology and tissue-specific expression profiles of V. officinalis putative P450s led to the discovery of a V. officinalis valerena-4,7(11)-diene oxidase, VoCYP71DJ1, which required coexpression with a V. officinalis alcohol dehydrogenase and aldehyde dehydrogenase to complete valerenic acid biosynthesis in yeast. Further, we demonstrated the stable integration of all pathway enzymes in yeast, resulting in the production of 140 mg/L of valerena-4,7(11)-diene and 4 mg/L of valerenic acid in milliliter plates. These findings showcase Saccharomyces cerevisiae's potential as an expression platform for facilitating multiply-oxidized medicinal terpenoid pathway discovery, possibly paving the way for scale up and FDA approval of valerenic acid and other active compounds from plant-derived herbal medicines.

The aim of this study was to investigate the beneficial effect of Valerian/Cascade mixture on sleeping in mammal models. In pentobarbital-induced sleep model, Valerian, Cascade, and Valerian/Cascade mixture significantly reduced the latency time for sleeping, and total sleeping time effectively increased in these sample groups compared with the control. Valerian/Cascade mixture increased sleep duration by 37%. The mixture significantly increased the non-rapid eye movement (NREM) sleep time by 53% compared with the control, while REM sleeping time was decreased by 33% with Valerian/Cascade mixture, in Electroencephalography (EEG) analysis, resulting in the increase of total sleep time and the decrease of awakening. This sleep-promoting effect was obvious in caffeine-induced awakening model; Valerian, Cascade, and the mixture significantly enhanced NREM and total sleep time, which were reduced by caffeine. Caffeine-induced increase of awakening was effectively deceased to the normal level by these three samples. In particular, delta wave responsible for deep sleep in NREM was greatly increased by the mixture in both normal and caffeine-induced awake models. This sleep-promoting effect of Valerian/Cascade mixture was shown to be due to the upregulation of gamma-aminobutyric acid A receptor (GABAAR). Valerian/Cascade mixture showed 91% binding capacity to GABAA-BZD receptor. Two compounds, Valerenic acid and Xanthohumol, were shown to significantly contribute to the binding activity of Valerian/Cascade mixture on the GABA receptor.

Valeriana officinalis is a medicinal herb which produces a suite of compounds in its root tissue useful for treatment of anxiety and insomnia. The sesquiterpene components of the root extract, valerenic acid and valerena-1,10-diene, are thought to contribute to most of the observed anxiolytic of Valerian root preparations. However, valerenic acid and its biosynthetic intermediates are only produced in low quantities in the roots of V. officinalis. Thus, in this report, Escherichia coli was metabolically engineered to produce substantial quantities of valerena-1,10-diene in shake flask fermentations with decane overlay. Expression of the wildtype valerenadiene synthase gene (pZE-wvds) resulted in production of 12 μg/mL in LB cultures using endogenous FPP metabolism. Expression of a codon-optimized version of the valerenadiene synthase gene (pZE-cvds) resulted in 3-fold higher titers of valerenadiene (32 μg/mL). Co-expression of pZE-cvds with an engineered methyl erythritol phosphate (MEP) pathway improved valerenadiene titers 65-fold to 2.09 mg/L valerenadiene. Optimization of the fermentation medium to include glycerol supplementation enhanced yields by another 5.5-fold (11.0 mg/L valerenadiene). The highest production of valerenadiene resulted from engineering the codon-optimized valerenadiene synthase gene under strong Ptrc and PT7 promoters and via co-expression of an exogenous mevalonate (MVA) pathway. These efforts resulted in an E. coli production strain that produced 62.0 mg/L valerenadiene (19.4 mg/L/OD600 specific productivity). This E. coli production platform will serve as the foundation for the synthesis of novel valerenic acid analogues potentially useful for the treatment of anxiety disorders.

Ferula sumbul (Umbelliferae) roots have a characteristic musk odour and it has been traditionally used for relieving anxiety, as a sedative in hysteria and other nervous disorders. Present investigation aims to develop and validate a simple, robust, accurate and fast analytical ultra-performance liquid chromatography technique for the estimation of valerenic acid in F. sumbul roots. F. sumbul root extract was prepared by refluxing the powdered roots with methanol for 30 minutes. A simple method for the estimation of valerenic acid has been developed and validated using ultra-performance liquid chromatography with photodiode array detector. The estimation was achieved using BEH Shield RP, C-18 column (2.1×50 mm, 1.7 μm) with isocratic elution of methanol and 0.5% phosphoric acid (75:25) at a flow rate of 0.25 ml/min and detection at 218 nm. The method has been developed for the first time and is accurate with good linearity (r2>0.998). F. sumbul roots were found to contain 12.62±0.074 µg/g valerenic acid. The developed method can be selectively used for the estimation of valerenic acid in the roots of F. sumbul. All the validation parameters were found to be within the permissible limits prescribed by ICH guidelines.

Valerenic acid (VA) is a pharmacologically-active sesquiterpene found in valerian (Valeriana officinalis L., Valerianaceae) roots and rhizomes. The plant produces only small amounts of this metabolite naturally. So, induction of hairy roots as well as elicitation can be useful to increase its commercial production. In this study, Wild-type strain ‘A13’ of Agrobacterium rhizogenes was used to induce hairy roots in valerian. The influence of three different elicitors including Fusarium graminearum extract (FE), methyl jasmonate (MJ) and salicylic acid (SA) on VA production in the selected hairy root line ‘LeVa-C4’ was also investigated. The 23-day-old cultures were treated with different concentrations of the elicitors at exposure time of 3 and 7 days. FE (1%) and MJ (100 µM L−1) highly promoted VA production at 7 days after elicitation, to a level of 12.31- and 6-fold higher than that of non-elicited controls, respectively, and FE did not exert any negative effects on biomass yield of hairy root. SA did not significantly increase the production of VA. This is the first time study to assess the elicitation of hairy root cultures to promote VA biosynthesis in valerian and the resulting experiments demonstrated that F. graminearum extract and MJ were indeed a potent inducer of VA biosynthesis.

Valerenic acid (VA) is a β2/3 subunit-specific modulator of γ-aminobutyric acid (GABA) type A (GABAA) receptors inducing anxiolysis. Here we analyze if VA-esters can serve as prodrugs and if different ester structures have different in vitro/in vivo effects. Modulation of GABAA receptors expressed in Xenopus oocytes was studied with 2-microelectrode-voltage-clamp. Anxiolytic effects of the VA-esters were studied on male C57BL/6N mice by means of the elevated plus maze-test; anticonvulsant properties were deduced from changes in seizure threshold upon pentylenetetrazole infusion. VA was detected in plasma confirming hydrolysis of the esters and release of VA in vivo. Esterification significantly reduced the positive allosteric modulation of GABAA (α1β3γ2S) receptors in vitro. in vivo, the studied VA-ester derivatives induced similar or even stronger anxiolytic and anticonvulsant action than VA. While methylation and propylation of VA resulted in faster onset of anxiolysis, the action of VA-ethylester was longer lasting, but occurred with a significant delay. The later finding is in line with the longer lasting anticonvulsant effects of this compound. The estimated VA plasma concentrations provided first insight into the release kinetics from different VA-esters. This might be an important step for its future clinical application as a potential non-sedative anxiolytic and anticonvulsant.

Valeriana officinalis is used in herbal medicine of many cultures as mild sedatives and tranquilizers. In this study, we investigated the effects of extract from valerian root extracts and its major component, valerenic acid on memory function, cell proliferation, neuroblast differentiation, serum corticosterone, and lipid peroxidation in adult and aged mice. For the aging model, D-galactose (100 mg/kg) was administered subcutaneously to 6-week-old male mice for 10 weeks. At 13 weeks of age, valerian root extracts (100 mg/kg) or valerenic acid (340 μg/kg) was administered orally to control and D-galactose-treated mice for 3 weeks. The dosage of valerenic acid (340 μg/kg), which is the active ingredient of valerian root extract, was determined by the content of valerenic acid in valerian root extract (3.401±0.066 mg/g) measured by HPLC. The administration of valerian root extract and valerenic acid significantly improved the preferential exploration of new objects in novel object recognition test and the escape latency, swimming speeds, platform crossings, and spatial preference for the target quadrant in Morris water maze test compared to the D-galactose-treated mice. Cell proliferation and neuroblast differentiation were significantly decreased, while serum corticosterone level and lipid peroxidation in hippocampus were significantly increased in the D-galactose-treated group compared to that in the control group. The administration of valerian root extract significantly ameliorated these changes in the dentate gyrus of both control and D-galactose-treated groups. In addition, valerenic acid also mitigated the D-galactose-induced reduction of these changes. These results indicate that valerian root extract and valerenic acid enhance cognitive function, promote cell proliferation and neuroblast differentiation, and reduce serum corticosterone and lipid peroxidation in aged mice.

major inhibitory GABA systems are known to take part a vital role in epilepsy, associated with excessive neuronal circuitry excitation. This excitatory action reflects seizures which result from GABA inhibitory circuit dysfunction. GABA mediates its fast inhibition through GABAA receptors by activating it and then opens the chloride channels. This opening allows chloride ions to flow into the interior of the cell which inhibits the excitability. This inhibitory action devises anticonvulsive properties as a consequence. So GABAA receptors are primary targets in the pathophysiology of epilepsy. Enhancement in the the action of GABAA receptors is the basis of epileptic seizures reduction on which many antiseizure drugs act. Modulation of GABAA receptor can be shown by many synthetic and natural compounds. Valerenic acid, a plant origin compound shows modulatory effects on GABAA receptors. In our present study, the homology model of the GABAA receptor subunit beta-3 is build and docking studies are performed with Valerenic acid. Docking analysis is done to know the binding interactions and their binding affinity. The results revealed the interacting amino acids which are involved in binding of GABAA receptor subunit beta-3 with Valerenic acid are ASP35, GLN56, PHE55, TYR120, ARG111 and TYR 54.

Made available in DSpace on 2018-11-26T15:29:25Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-05-02 Universidade Estadual do Sudoeste da Bahia Pontificia Universidad Javeriana Jovenes Investigadores (Colciencias) Parkinson's disease (PD) is one of the most common neurodegenerative disorders. A theoretical approach of our previous experiments reporting the cytoprotective effects of the Valeriana officinalis compounds extract for PD is suggested. In addiction to considering the PD as a result of mitochondrial metabolic imbalance and oxidative stress, such as in our previous in vitro model of rotenone, in the present manuscript we added a genomic approach to evaluate the possible underlying mechanisms of the effect of the plant extract. Microarray of substantia nigra (SN) genome obtained from Allen Brain Institute was analyzed using gene set enrichment analysis to build a network of hub genes implicated in PD. Proteins transcribed from hub genes and their ligands selected by search ensemble approach algorithm were subjected to molecular docking studies, as well as 20 ns Molecular Dynamics (MD) using a Molecular Mechanic Poison/Boltzman Surface Area (MMPBSA) protocol. Our results bring a new approach to Valeriana officinalis extract, and suggest that hesperidin, and probably linarin are able to relieve effects of oxidative stress during ATP depletion due to its ability to binding SUR1. In addition, the key role of valerenic acid and apigenin is possibly related to prevent cortical hyperexcitation by inducing neuronal cells from SN to release GABA on brain stem. Thus, under hyperexcitability, oxidative stress, asphyxia and/or ATP depletion, Valeriana officinalis may trigger different mechanisms to provide neuronal cell protection. Univ Estadual Sudoeste Bahia, Dept Ciencias Biol, Jequie, Brazil Univ Estadual Sudoeste Bahia, Dept Quim & Exatas, PNPD CAPES, Jequie, Brazil Pontificia Univ Javeriana, Fac Ciencias, Dept Nutr & Bioquim, Bogota, DC, Colombia UBA CONICET, Inst Invest Cardiol Prof Dr Alberto C Taquini INI, Buenos Aires, DF, Argentina Univ Estadual Paulista, Inst Biol, Dept Bioquim, Campinas, SP, Brazil Univ Fed Estadual Feira Santana, Dept Ciencias Biol, Feira De Santana, Brazil Univ Autonoma Chile, Inst Ciencias Biomed, Santiago, Chile Univ Cient Sur, Lima, Peru Univ Estadual Paulista, Inst Biol, Dept Bioquim, Campinas, SP, Brazil Pontificia Universidad Javeriana: 5575 Pontificia Universidad Javeriana: 5024 Jovenes Investigadores (Colciencias): 6027

Valeriana fauriei (V. fauriei), which emits a characteristic and unpleasant odor, is important in traditional medicine. In this study, the expression of terpenoid biosynthetic genes was investigated in different organs that were also screened for volatile compounds including valerenic acid and its derivatives. Specific expression patterns from different parts of V. fauriei were observed using quantitative real-time PCR (qRT-PCR). The highest transcript levels of biosynthetic genes involved in mevalonic acid (MVA) and methylerythritol phosphate (MEP) production were found in the stem. Although the amounts of volatile compounds were varied by organ, most of the volatile terpenoids were accumulated in the root. Gas chromatography mass spectrometry (GC-MS) analysis identified 128 volatile compounds, which represented 65.33% to 95.66% of total volatiles. Certain compounds were only found in specific organs. For example, isovalerenic acid and valerenic acid and its derivatives were restricted to the root. Organs with high transcript levels did not necessarily have high levels of the corresponding chemical constituents. According to these results, we hypothesize that translocation may occur between different organs in V. fauriei.

Ziel dieser Arbeit war die systematische Untersuchung von GABAA-Rezeptor Modulatoren, welche strukturell auf dem Naturstoff Valerens��ure basieren. Hierzu wurden drei verschiedene strukturelle Eigenschaften - durch chemische Modifikation und anschlie��ender pharmakologischer Untersuchung- variiert, um deren Einfluss auf Effektivit��t und Selektivit��t am GABAA-Rezeptor zu determinieren. Folglich stellt jede Modifikation ein Kapitel dieser Arbeit dar. Das erste Kapitel befasst sich mit der Derivatisierung der Carboxylat-Funktion. Hier galt es herauszufinden, durch welche chemischen Eigenschaften die einzig polare (heteroatomare) Gruppe ihre Wirkung am Rezeptor entfaltet. Diesbez��glich wurde diese Funktionalit��t durch Derivatisierung in verschiedene Ester und Amide vor allem auf die Notwendigkeit von Wasserstoffbr��ckenakzeptor- beziehungsweise Wasserstoffbr��ckendonor-Wechselwirkungen untersucht. Aus den 17 verschiedenen synthetisierten S��urederivaten (inklusive dem Bioisoster Tetrazol) konnte bewiesen werden, dass eine Wasserstoffbr��ckendonor-Eigenschaft f��r eine pharmakologisch relevante Wirkung unabdingbar ist. Weiters wurden noch Untersuchungen - und damit chemische Modifikationen - an beiden exocyclischen Methylgruppen der Indanylkernstruktur durchgef��hrt. Auch hier galt als Zielsetzung die Notwendigkeit beider Methylsubstituenten f��r GABAA- Subtypenselektivit��t und Effektivit��t herauszufinden. Dies konnte durch Substitution der entsprechenden Methylgruppen durch unterschiedlich gro��e Alkyl-respektive Alkenylreste demonstriert werden. Als Ergebnis dieser Untersuchung sei hier vor allem die entscheidende Rolle des axialen Methylsubstituenten in 7-Position hervorzuheben, da jegliche Modifikation der Methylgruppe in einem drastischen Verlust an Effektivit��t und Subtypenselektivit��t resultiert. Des Weiteren konnte gezeigt werden, dass f��r den vinylogen Substituent in 3-Position nur eine limitierte r��umliche Ausdehnung (maximal Ethyl) toleriert wird. Schlie��lich wurden auch synthetische Limitierungen der verwendeten, publizierten Valerens��ure-Totalsynthese bez��glich ihrer Modulation in 7-Position aufgezeigt. Infolgedessen konnte dieser Synthesezugang leicht abgewandelt werden und stellt somit einen Zugang f��r bisher weder bekannte, noch auf diesem Weg machbare, Valerens��uremodifikationen dar. Kurzum stellt der in dieser Arbeit entwickelte Syntheseweg sowohl eine kosteng��nstigere als auch bei weitem modularere Alternative dar., The main goal of this Thesis was the investigation of GABAA-receptor modulators, based on the natural product valerenic acid. Therefore, variation of three different structural features should - through chemical modification and subsequent pharmacological investigation- determine the influence of these modifications on efficacy and selectivity. Each modification made, represents a chapter within this thesis. The first chapter contains the derivatization of the carboxylic acid function. It was crucial to study the pharmacological effects exhibited by the chemical properties of the only polar (heteroatomic) group. Therefore, this functionality was transformed into various esters and amides, to investigate the role of hydrogen acceptor or hydrogendonor interactions. Through 17 synthesized acid-derivatives (including the carboxylic acid bioisostere tetrazole) it could be proven, that a hydrogen donor interaction is crucial for the observed pharmacological effect. Furthermore the role of both exocyclic methyl substituents, attached on the indanyl core, was investigated in terms of efficacy and selectivity. This was accomplished through appropriate substitution of each methyl group via alkyl- respectively alkenyl residues of different size. As a result, methyl substitution in 7-position seems to be crucial for both efficacy and selectivity, as each modification led to severe loss of both pharmacological parameters. Furthermore, it could be demonstrated, that at maximum an ethyl substituent is tolerated for the vinylogous substituent in 3-position, as each larger modification significantly lowered efficacy and selectivity at the GABAA-receptor. Finally, synthetic limitaions of the published valerenic acid total synthesis in terms of modularity could be illustrated. Therefore, a slightly different synthetic approach was developed to enable structural variations - especially in 7-position- yet not accessible through published syntheses. As a result, a more economical and versatile alternative synthesis for valerenic acid derivatives was developed within this thesis.

Valerian is one of the most commonly used herbal remedies for the treatment of insomnia and anxiety. Valerian extracts allosterically modulate GABAA receptors, an action related to valerenic acid, which is one of the active compounds determined from pharmacological studies. Derivatives of valerenic acid, i.e. acetoxy valerenic acid or hydroxy valerenic acid, do not allosterically modulate GABAA receptors, but they bind to identical binding sites. Therefore, the question arises whether they might interfere with the effects of valerenic acid. Two valerian extracts were tested in the elevated plus maze test and the tail suspension test for anxiolytic and antidepressive activity, respectively. Reference substances were diazepam (1.0mg/kg) and imipramine (30mg/kg). The extracts were standardized to the identical total amounts of the acids (0.1; 0.5; 1.0 and 2.0mg/kg), i.e. valerenic and acetoxy valerenic acid, but the ratio between the acids was different (12:1 and 1:1.5). The extract with the ratio 12:1 prolonged the time spent on the open arm significantly when 0.5mg/kg was applied. Of the other extract, with the ratio 1:1.5, four times that amount was required (2.0mg/kg). Both of the tested extracts did not show any antidepressive effect, rather the other way around, the extract with the ratio 1:1.5 prolonged the immobility phase. However, since the core body temperature was reduced by the 1.0 and 2.0mg/kg extract dose, the prolongation may be related to the temperature phenomenon and is not indicative of a specific depressive action. In conclusion, the anxiolytic activity of the valerian extract seems rather related to valerenic acid and, moreover, standardization with respect to the total amount of valerenic acids, i.e. valerenic acid together with acetoxy valerenic acid, is misleading.

Valerian extract is used in complementary and alternative medicine for its anxiolytic and sedative properties. Our previous research demonstrated valerian interactions with glutamate receptors. The purpose of this study was to determine if valerian anxiolytic properties are mediated by metabotropic glutamate receptors (mGluR) such as mGluR (1/5) (mGluR I) and mGluR (2/3) (mGluR II). Adult wild-type zebrafish (Danio rerio) prefer the black compartment and avoid the white compartment in the dark/light preference task. Zebrafish exposed to 1 mg/mL of valerian extract or 0.00117 mg/mL valerenic acid increased their residence time in the white side by 84.61 ± 6.55 % and 58.30 ± 8.97 %, respectively. LAP3 (mGluR I antagonist) and EGLU (mGluR II antagonist) significantly inhibited the effects of valerian and valerenic acid. These results demonstrated that valerian and valerenic acid have anxiolytic properties in the zebrafish. Moreover, the selective interaction of valerian with mGluR I and II represent an alternative explanation for the anxiolytic properties of this plant and support the role of mGluR in anxiety.

Valerian (Valeriana officinalis) is a popular medicinal plant in North America and Europe. Its root extract is commonly used as a mild sedative and anxiolytic. Among dozens of chemical constituents (e.g. alkaloids, iridoids, flavonoids, and terpenoids) found in valerian root, valerena-4,7(11)-diene and valerenic acid (C15 sesquiterpenoid) have been suggested as the active ingredients responsible for the sedative effect. However, the biosynthesis of the valerena-4,7(11)-diene hydrocarbon skeleton in valerian remains unknown to date. To identify the responsible terpene synthase, next-generation sequencing (Roche 454 pyrosequencing) was used to generate ∼ 1 million transcript reads from valerian root. From the assembled transcripts, two sesquiterpene synthases were identified (VoTPS1 and VoTPS2), both of which showed predominant expression patterns in root. Transgenic yeast expressing VoTPS1 and VoTPS2 produced germacrene C/germacrene D and valerena-4,7(11)-diene, respectively, as major terpene products. Purified VoTPS1 and VoTPS2 recombinant enzymes confirmed these activities in vitro, with competent kinetic properties (K(m) of ∼ 10 μm and k(cat) of 0.01 s(-1) for both enzymes). The structure of the valerena-4,7(11)-diene produced from the yeast expressing VoTPS2 was further substantiated by (13) C-NMR and GC-MS in comparison with the synthetic standard. This study demonstrates an integrative approach involving next-generation sequencing and metabolically engineered microbes to expand our knowledge of terpenoid diversity in medicinal plants.

A detailed study of the total synthesis of valerenic acid, a well known GABAA receptor subtype modulator, is described. Both successful as well as unsuccessful attempts towards the synthesis of the title compound are presented, including four different strategies to synthesize one of the key intermediates. The first two strategies are based on epoxides provided from the chiral pool, whereas the last two approaches rest on stereocontrolled modifications of 2-cyclopentenone. The streamlined synthesis implements a new one-pot reaction, which combines the addition of a Grignard species with an acid-catalyzed isomerization of the intermediate allylic alcohol. Further highlights are a stereo- and regioselective hydroxy-directed Diels–Alder reaction, a hydroxy-directed hydrogenation, and a final Negishi coupling reaction. After optimization of our synthesis, the preparation of several easily available derivatives is also discussed. Amides obtained by functionalization of the carboxyl group are more than twice as active as valerenic acid.

The changes in total phenolics, flavonoids, tannins, valerenic acid, and antioxidant activity were assessed in 25 populations of Valeriana jatamansi sampled from 1200 to 2775 m asl and four habitat types of Uttarakhand, West Himalaya. Significant (p

In the present work, the applicability of hollow fiber-based liquid phase microextraction (HF-LPME) was evaluated for the extraction and preconcentration of valerenic acid prior to its determination by reversed-phase HPLC/UV. The target drug was extracted from 5.0 mL of aqueous solution with pH 3.5 into an organic extracting solvent (dihexyl ether) impregnated in the pores of a hollow fiber and finally back extracted into 10 μ L of aqueous solution with pH 9.5 located inside the lumen of the hollow fiber. In order to obtain high extraction efficiency, the parameters affecting the HF-LPME, including pH of the donor and acceptor phases, type of organic phase, ionic strength, the volume ratio of donor to acceptor phase, stirring rate and extraction time were studied and optimized. Under the optimized conditions, enrichment factor up to 446 was achieved and the relative standard deviation (RSD) of the method was 4.36% (n = 9). The linear range was 7.5-850 μg L⁻¹ with correlation coefficient (r²=0.999), detection limits was 2.5 μg L⁻¹ and the LOQ was 7.5 μg L⁻¹. The proposed method was evaluated by extraction and determination of valerenic acid in some Iranian wild species of Valerianaceae.

Although GABA neurotransmission has been suggested as a mechanism forValeriana officinaliseffects, CNS depression can also be evoked by inhibition of ionotropic (iGluR) and metabotropic glutamate receptors (mGluR). In this study, we examined if aqueous valerian extract interacted with glutamatergic receptors. Freshly prepared aqueous valerian extract was incubated with rat cortical synaptic membranes in presence of 20 nM [3H]Glutamate. Aqueous valerian extract increased [3H]Glutamate binding from1×10−7to1×10−3 mg/mL. In the presence of (2S,1′S,2′S)-2-(Carboxycyclopropyl)glycine (LCCG-I) and (2S,2′R,3′R)-2-(2′,3′-Dicarboxycyclopropyl)glycine (DCG-IV), Group II mGluR agents, valerian extract markedly decreased [3H]Glutamate binding, while (2S)-2-amino-3-(3,5-dioxo-1,2,4-oxadiazolidin-2-yl) propanoic acid) (quisqualic acid, QA), Group I mGluR agonist, increased [3H]Glutamate binding. At 0.05 mg/mL aqueous valerian extract specifically interacted with kainic acid NMDA and AMPA receptors. Valerenic acid, a marker compound forValeriana officinalis, increased the [3H]Glutamate binding after1.6×10−2 mg/mL, and at 0.008 mg/mL it interacted only with QA (Group I mGluR). The selective interactions of valerian extract and valerenic acid with Group I and Group II mGluR may represent an alternative explanation for the anxiolytic properties of this plant.

BACKGROUND AND PURPOSE Subunit-specific modulators of γ-aminobutyric acid (GABA) type A (GABAA) receptors can help to assess the physiological function of receptors with different subunit composition and also provide the basis for the development of new drugs. Valerenic acid (VA) was recently identified as a β2/3 subunit-specific modulator of GABAA receptors with anxiolytic potential. The aim of the present study was to generate VA derivatives as novel GABAA receptor modulators and to gain insight into the structure–activity relation of this molecule. EXPERIMENTAL APPROACH The carboxyl group of VA was substituted by an uncharged amide or amides with different chain length. Modulation of GABAA receptors composed of different subunit compositions by the VA derivatives was studied in Xenopus oocytes by means of the two-microelectrode voltage-clamp technique. Half-maximal stimulation of GABA-induced chloride currents (IGABA) through GABAA receptors (EC50) and efficacies (maximal stimulation of IGABA) were estimated. Anxiolytic activity of the VA derivatives was studied in mice, applying the elevated plus maze test. KEY RESULTS Valerenic acid amide (VA-A) displayed the highest efficacy (more than twofold greater IGABA enhancement than VA) and highest potency (EC50= 13.7 ± 2.3 µM) on α1β3 receptors. Higher efficacy and potency of VA-A were also observed on α1β2γ2s and α3β3γ2s receptors. Anxiolytic effects were most pronounced for VA-A. CONCLUSIONS AND IMPLICATIONS Valerenic acid derivatives with higher efficacy and affinity can be generated. Greater in vitro action of the amide derivative correlated with a more pronounced anxiolytic effect in vivo. The data give further confidence in targeting β3 subunit containing GABAA receptors for development of anxiolytics.

Valerian root (Valeriana officinalis) is a popular and widely available herbal supplement, primarily used to treat insomnia and anxiety. Until recently, its mechanism of action has remained unknown. Neurobiological research has begun to show that the herb, with its active valerenic acid, interacts with the GABA(A)-ergic system, a mechanism of action similar to the benzodiazepine drugs. This series of experiments sought to corroborate these findings with behavioral measures, compare them to the benzodiazepine diazepam, and to analyze the chemical composition of Valeriana officinalis. Rats were administered either ethanol (1 ml/kg), diazepam (1mg/kg), valerian root extract (3 ml/kg), valerenic acid (3mg/kg), or a solution of valerenic acid and exogenous GABA (75 microg/kg and 3.6 microg/kg, respectively) and assessed for the number of entries and time spent on the open arms of an elevated plus maze. Results showed that there was a significant reduction in anxious behavior when valerian extract or valerenic acid exposed subjects were compared to the ethanol control group. The evidence supports Valeriana officinalis as a potential alternative to the traditional anxiolytics as measured by the elevated plus maze.

Supercritical CO2 extraction from valerian (Valeriana officinalis L.) roots was studied. The objective of this study was to optimize the independent process parameters (temperature, pressure, dynamic time and modifier volume) in order to attain the highest yield of the essential oil and valerenic acids as the desired compounds. The process optimization was based on a simplex centroid design (mixture design), coupled with the statistical and graphical analysis of the results. Due to the features of mixture method and to the proper ratios of four independent variables, the optimal points were in the range of temperature (37 °C), pressure (24.3–25.0 MPa), dynamic time (19–24 min) and modifier volume (100–200 μl).

The genus Valeriana (Valerianaceae) contains over 250 species distributed around the world and used in traditional medicines of many cultures. The main compounds isolated from Valeriana species are essential oils, valerenic acid and its derivatives and iridoids, accumulated mainly in the roots and rhizomes. This review lists 135 chemical constituents as well as their biosynthesis and bioactivity as reported by the end of 2008 (80 references).

Insomnia is a commonly reported clinical problem with as many as 50% of older adults reporting difficulty in falling and/or remaining asleep. Valerian (Valeriana officinalis) is a commonly used herb that has been advocated for promoting sleep. Valerenic acid is used as a marker for quantitative analysis of valerian products with evidence of pharmacological activity relevant to the hypnotic effects of valerian. The objective of this study was to determine the pharmacokinetics of valerenic acid in a group of elderly women after receiving a single nightly valerian dose and after 2 weeks of valerian dosing. There was not a statistically significant difference in the average peak concentration (C(max)), time to maximum concentration (T(max)) area under the time curve (AUC), elimination half-life (T(1/2)) and oral clearance after a single dose compared with multiple dosing. There was considerable inter- and intra-subject variability in the pharmacokinetic parameters. C(max) and AUC deceased and T(1/2) increased with increased body weight. The variability between the capsules was extremely low: 2.2%, 1.4% and 1.4%, for hydroxyvalerenic acid, acetoxyvalerenic acid and valerenic acid, respectively. In conclusion, large variability in the pharmacokinetics of valerenic acid may contribute to the inconsistencies in the effect of valerian as a sleep aid.

The study was designed to investigate the hepatic metabolism and transport system of valerenic acid, a main active constituent of valerian, in isolated perfused livers from Wistar and Mrp2-deficient TR(-) rats. After administration of 20 microM valerenic acid, the formation of seven valerenic acid glucuronides (M1-M7), namely two glucuronides of valerenic acid (M6, M7), four glucuronides of hydroxylated valerenic acid (M1, M3, M4, M5), and one glucuronide of hydroxylated dehydro-valerenic acid (M2) in bile and perfusate was quantified by HPLC. The hepatic extraction ratio and clearance of valerenic acid were very high in Wistar and TR(-) rats (E: 0.983 +/- 0.006 vs. 0.981 +/- 0.004; Cl: 35.4 +/- 0.21 mL/min vs. 35.3 +/- 0.14 mL/min). However, biliary excretion and efflux of conjugates differed greatly in TR(-) rats. While cumulative biliary excretion of unconjugated valerenic acid and the glucuronides M1-M7 dropped dramatically to 1-9%, their efflux into perfusate increased 1.5- to 10-fold. This indicates that valerenic acid and its glucuronides are eliminated into bile by Mrp2. In summary, valerenic acid was metabolized to several conjugates, whereby the canalicular transporter Mrp2 mediated biliary excretion of the parent drug and its glucuronides.

The content of essential oil and valerenic acids and the yield of roots were analyzed in valerian cultivar ‘Polka’ and strain no. 4 during the nine following phases: the beginning of vegetation, spring rosette, pedicel development, the beginning of flowering, full bloom, seed harvest, three weeks after the seed harvest, fall rosette, and after the first frost. The yield of valerian roots was different at the investigated phases and in years of cultivation, while tested genotypes produced similar root yields. The highest contents of essential oil and valerenic acids were obtained at different phases by the tested genotypes: strain no. 4 in full bloom (1.3% and 0.266%, respectively) and cultivar ‘Polka’ three weeks after seed harvest (1.25%) and during seed harvest (0.218%). The lowest contents of the investigated substances were detected at the beginning of growth and after the first frost in both genotypes. The obtained results revealed significant differences in essential oil and valerenic acids...

Dried valerian ( Valeriana officinalis ) root powder was stored at 5, 14 and 30 °C under low, moderate and high humidity conditions for 6 months, and the level of the valerenic acids monitored on a monthly basis. From an initial moisture content of 5 g/kg, the moisture level during storage decreased in root powder stored at low humidity and increased during storage at high humidity, with the effect greatest at 30 °C. The concentration of valerenic acid significantly decreased over time and was affected by temperature and humidity with the loss greatest at 30 °C in low humidity. Acetoxyvalerenic acid also decreased over time but with greatest loss at 30 °C and high humidity. Hydroxyvalerenic acid was not detected in the fresh sample but was present in all stored samples and tended to reflect the decrease in acetoxyvalerenic acid. The concentration of total valerenic acids essentially mirrored the change in valerenic acid. It is concluded that maximum retention of valerenic acids would be achieved by storing valerian at a temperature lower than about 14 °C with humidity not a great concern. If storage is to occur at higher temperatures, retention of valerenic acids is favoured by storage under a high humidity but such a recommendation is not considered practical due to the risk of microbial growth and increasing difficulty of postharvest handling of moist material.

Aim To study the chemical constituents of the roots of Valeriana officinalis L. Methods Chemical constituents were isolated by various column chromatographic techniques and their structures were elucidated by spectroscopic methods. Results A new sesquiterpene, orivalerianol (4β, 10α, 15-trihydroxy-aromadrendane) ( 1 ), and a new iridoid, monovalerianester A (aglycone of kanokoside A) ( 2 ) together with eight known compounds rulepidol ( 3 ), behenic acid ( 4 ), pinoresinol ( 5 ), valerenic acid ( 6 ), β-sitosterol ( 7 ), kanokoside A ( 8 ), prinsepiol-4- O -β- D -glucoside ( 9 ) and 8-hydroxypinoresinol-4 - O -β- D -glucoside ( 10 ) were obtained from the EtOAc extract ( 1∼6 ) and n -BuOH extract ( 7∼10 ). Conclusion Compounds 1 and 2 are new natural products.

Valerenic acid is a major constituent of valerian root. It exhibits modulatory activity on the GABA A receptor and thus represents a potential new lead structure for the discovery of new anxiolytics. Here we present the enantioselective total synthesis of valerenic acid, which departs from natural (R)-pulegone and proceeds through a bicyclic ketone as the central intermediate. Key transformations are the stereoselective reduction of a 3,4-disubstituted cyclohexenone and a microwave-assisted Wittig reaction.

Valerian extracts have been used for centuries to alleviate restlessness and anxiety albeit with unknown mechanism of action in vivo. We now describe a specific binding site on GABA(A) receptors with nM affinity for valerenic acid and valerenol, common constituents of valerian. Both agents enhanced the response to GABA at multiple types of recombinant GABA(A) receptors. A point mutation in the beta2 or beta3 subunit (N265M) of recombinant receptors strongly reduced the drug response. In vivo, valerenic acid and valerenol exerted anxiolytic activity with high potencies in the elevated plus maze and the light/dark choice test in wild type mice. In beta3 (N265M) point-mutated mice the anxiolytic activity of valerenic acid was absent. Thus, neurons expressing beta3 containing GABA(A) receptors are a major cellular substrate for the anxiolytic action of valerian extracts.

One of the important active compounds of valerian is valerenic acid, which is supposed to be the chemical factor responsible for the sedative effect of valerian. The aim of this work is identificat...

The dynamic behavior of supercritical fluid extraction (SFE) of valerenic acid (VA) from valerian (Valeriana officinalis L.) roots was studied by mathematical modeling. The extraction yield of VA was considered as the most desirable compound among the other extracted constituents. A two-phase desorption model was developed by considering a diffusion controlled regime in the particle and axial dispersion in the bed. The mass transfer parameters, i.e., pore diffusivity, film mass transfer coefficient and axial dispersion, along with the solubility parameters were chosen as the model parameters. The first three mass transfer parameters were predicted using nondimensional equations from the literature. The solubility equation and the parameters were studied using different equilibrium models, i.e., Henry, Langmuir, Freundlich, Langmuir-Freundlich (L-F) and Toth isotherms. The equilibrium parameters were correlated by comparing the outlet results of the dynamic SFE model with experiments. The experimental yield of the VA extraction was obtained at a pressure of 15.0–36.0 MPa, temperature of 310–334 K, solvent flow rate of 0.50–1.10 · 10–6 m3/min and different particle sizes ranging from 0.18–2.00 · 10–3 m in diameter, at a 20 min constant static period, in the presence of 46.9 μL/g ethanol as the co-solvent, followed by dynamic time extraction for up to 50 min. From the results, the mathematical model using the L-F equation exhibited the best agreement with the experimental yield of VA extraction in the range of studied conditions. The present model can be applied to design and scale up the SFE process of VA from Valeriana officinalis L. roots.

An approach for rapid HPLC-based profiling for new GABA (A) ligands of natural origin has been developed. Active extracts are separated by a single injection of 3-10 mg of extract onto a semi-preparative (150 x 10 mm i. d.) HPLC column with gradient elution and time-based fractionation. The microfractions are tested in an automated two-microelectrode voltage-clamp assay on Xenopus oocytes expressing recombinant GABA (A) channels composed of alpha (1), beta (2) and gamma (2S) subunits. The protocol has been validated by spiking experiments with inactive extract and the GABA (A) receptor ligand magnolol, and by profiling of active extracts such as valerian extract containing the known GABA (A) receptor ligand valerenic acid. For dereplication of GABA containing extracts, we established a rapid and simple procedure by which GABA is analyzed as OPA derivative by reversed-phase HPLC. This dereplication protocol was validated with plant and fungal extracts which had been previously tested active or inactive in the oocyte assay and with spiking experiments.

The composition of essential oil extracted from Valeriana officinalis L. roots growing wild in Iran was studied by hydrodistillation and supercritical CO2 extraction. Forty-seven components representing 89.3% and 35 constituents varying from 86.1% to 95.1% of the oil obtained by hydrodistillation and supercritical CO2 were identified, respectively. The major components in the extracted oil from supercritical CO2 were isovaleric acid (18.7-41.8%), valerenic acid (8.2-11.8%), acetoxyvaleranone (5.6-9.6%), (Z)-valernyl acetate (4.5-6.5%), bornyl acetate (2.3-7.7%) and valerenol (3.7-5.2%), whereas by hydrodistillation were bornyl acetate (11.6%), valerenic acid (8.0%), (Z)-valernyl acetate (7.9%) and acetoxyvaleranone (7.6%). The analysis of the extracts was performed by capillary GC and GC/MS.

Valeriana Officinalis L . is a traditionally used sleep remedy, however, the mechanism of action and the substances responsible for its sedative and sleep-enhancing properties are not fully understood. As we previously identified valerenic acid as a subunit-specific allosteric modulator of GABAA receptors, we now investigated the relation between modulation of GABAA receptors by Valerian extracts of different polarity and the content of sesquiterpenic acids (valerenic acid, acetoxyvalerenic acid). All extracts were analysed by HPLC concerning the content of sesquiterpenic acids. GABAA receptors composed of alpha 1, beta 2 and gamma 2S subunits were expressed in Xenopus laevis oocytes and the modulation of chloride currents through GABAA receptors (IGABA) by Valerian extracts was investigated using the two-microelectrode voltage clamp technique. Apolar extracts induced a significant enhancement of IGABA, whereas polar extracts showed no effect. These results were confirmed by fractionating a highly active ethyl acetate extract: again fractions with high contents of valerenic acid exhibited strong receptor activation. In addition, removal of sesquiterpenic acids from the ethyl acetate extract led to a loss of I (GABA) enhancement. In conclusion, our data show that the extent of GABAA receptor modulation by Valerian extracts is related to the content of valerenic acid.

The pharmacological activity of neuroactive phytochemicals on recombinant human glutamic acid decarboxylase 65 (hGAD65) was investigated. GAD catalyzes the conversion of glutamic acid to γ-aminobutyric acid (GABA), which acts as an important inhibitory neurotransmitter in the central nervous system (CNS). We describe an improved method in which recombinant hGAD65 was expressed at high levels using a maltose binding protein (MBP) fusion system. The expression and purification process was superior to the commonly used glutathioneS.-transferase (GST) fusion partner. The in vitro. system developed here detected both enzyme inhibition and stimulation, under varying pyridoxal-5′-phosphate (PLP) concentrations. The known GAD inhibitor, 3-mercaptopropionic acid, was tested as a positive control and had an IC50 = 12.3 μ M. Phytochemicals were tested (10 μ g/mL) for their effects on in vitro. hGAD65 activity. Minor inhibition was seen with the ethanol extract of Panax quinquefolius. L. (ginsenosides) (23%),...

Background Anticonvulsant properties have been attributed to extracts of the herbal medicine Valeriana officinalis. Our aims were to examine the anticonvulsant properties of valerenic acid and valerian extracts and to determine whether valerian preparations interact with the activity of other anti-epileptic drugs (phenytoin or clonazepam). To achieve these goals, we validated the adult zebrafish, Danio rerio, as an animal model for studying anticonvulsant drugs. Methods All drug treatments were administered by immersion in water containing the drug. For assays of anticonvulsant activity, zebrafish were pretreated with: anti-epileptic drugs, valerenic acid, aqueous or ethanolic valerian extracts, or mixtures (phenytoin or clonazepam with valerenic acid or valerian extracts). Seizures were then induced with pentylenetetrazole (PTZ). A behavioral scale was developed for scoring PTZ-induced seizures in adult zebrafish. The seizure latency was evaluated for all pretreatments and control, untreated fish. Valerenic acid and both aqueous and ethanolic extracts of valerian root were also evaluated for their ability to improve survival after pentylenetetrazole-challenge. The assay was validated by comparison with well-studied anticonvulsant drugs (phenytoin, clonazepam, gabapentin and valproate). One-way ANOVA followed by Tukey post-hoc test was performed, using a p

Valeriana jatamansi Jones, commonly known as Indian Valerian or Tagar (Family: Valerinaceae) is used in both traditional and modern systems of medicine. The plants of this species are known for their high content of valerenic acid, the main active constituent of valerian, and high antioxidant activity, and these characteristics have increased the demand for these plants by the pharmaceutical industry. At present, the demand for planting material is largely met from the harvesting of wild plants, which vary in quality and quantity of the active ingredient. Therefore, there is a need to identify individual plants/populations with a higher content of the active ingredients and higher antioxidant activity. We used inter-simple sequence repeats (ISSR) markers in 151 genotypes of 25 V. jatamansi populations to identify markers associated with valerenic acid and antioxidant activity. Of the 130 ISSR primers tested, 20 generated 159 bands, with an average of 7.95 bands per primer. Valerenic acid content was significantly (p < 0.05) higher in the Katarmal (aerial portion 0.57 ± 0.04 %) and Joshimath populations (root portion 1.80 ± 0.12 %). Antioxidant activity using these different in vitro assays varied among the populations and plant portions, with maximum antioxidant activity found in the aerial plant portion (8.63 ± 0.06 mM) and roots [8.36 ± 0.0 mM ascorbic acid equivalents (AAE)/100 g dry weight (dw)] of the Didihat and Katarmal populations, respectively, using the ABTS [2, 2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic)] assay. The DPPH (2, 2-diphenyl-1-picryylhydrazyl free radical scavenging) assay revealed maximum antioxidant activity in the aerial plant portion (15.23 ± 0.09 mM) and roots (17.53 ± 0.04 mM AAE/100 g dw) of the Didihat population. The FRAP (ferric-reducing antioxidant power) assay showed that the roots of plants of the Ukhimath population had significantly higher antioxidant activity (12.71 ± 0.04 mM AAE/100 g dw) than those of other populations of V. jatamansi. Based on the stepwise multiple regression analysis, seven positive and six negative markers showed significant association with valerenic acid content. Antioxidant activity measured by the ABTS, DPPH and FRAP assays showed a positive correlation with 14, 13 and 10 markers, respectively (p < 0.001). These markers have the potential for application in breeding programmes in order to select lineages of V. jatamansi with higher biochemical attributes, especially when no other genetic information, such as linkage maps and quantitative trait loci is available.

Insomnia affects approximately one-third of the adult population and contributes to increased rates of absenteeism, health care use, and social disability. Extracts of the roots of valerian (Valeriana officinalis) are widely used for inducing sleep and improving sleep quality. A systematic review of randomized, placebo-controlled trials of valerian for improving sleep quality is presented. An extensive literature search identified 16 eligible studies examining a total of 1093 patients. Most studies had significant methodologic problems, and the valerian doses, preparations, and length of treatment varied considerably. A dichotomous outcome of sleep quality (improved or not) was reported by 6 studies and showed a statistically significant benefit (relative risk of improved sleep = 1.8, 95% confidence interval, 1.2-2.9), but there was evidence of publication bias in this summary measure. The available evidence suggests that valerian might improve sleep quality without producing side effects. Future studies should assess a range of doses of standardized preparations of valerian and include standard measures of sleep quality and safety.

A simple, rapid, cost-effective and accurate high performance thin layer chromatographic method has been developed for quantification of valerenic acid in Valeriana jatamansi and Valeriana officinalis which is one of the stable compounds of Valeriana officinalis and designated as a key marker compound. Valerenic acid makes substantial contribution to the sedative and spasmolytic activity of the essential oil and extract of Valeriana officinalis. Separation and quantification was achieved by HPTLC using ternary mobile phase of hexane: ethyl acetate: acetic acid (80:20:0.5 v/v) on precoated silica gel 60F254 aluminium plates and densitometric determination was carried out after derivatization with anisaldehyde-sulphuric acid reagent at 700 nm, in absorption-reflectance mode. The calibration curves were linear in the range of (500 ng–2.5 µg). This is the first HPTLC report for the identification and quantification of valerenic acid in Valeriana jatamansi and Valeriana officinalis.

Valeriana officinalis extracts are used in folkloric medicine for their sedative, hypnotic and tranquilizer effects. Using [3H]flunitrazepam binding as an indicator, the interactions of commercial Valerian extracts with GABAA receptors were examined. There was considerable fluctuation among the different extracts, some mildly enhanced [3H]flunitrazepam binding, others had no effect and others had inhibitory effects, independent of standardization by valerenic acid. Central depression can also be accomplished by a reduction of excitatory transmission. Valerian extracts had modest inhibitory effects on [3H]MK-801 binding, an indicator of NMDA–Valerian interactions. Spectral analyses (UV region) did not show marked differences among the different extracts. The inhibitory effects of one of the extracts on [3H]flunitrazepam binding was somewhat stable, while on [3H]MK-801 binding the inhibitory effects were lost within months. These results suggest that particular care should be taken in analysing and interpreting results from commercial Valerian preparations. Copyright © 2006 John Wiley & Sons, Ltd.

The chemical composition of essential oils obtained from fresh and dried roots of Valeriana officinalis L. (Valerianaceae) was analyzed by GC/FID and GC/MS using two columns of different polarities, polyethylene glycol (DB-Wax) and 5% phenyl 95% polydimethylsiloxane (HP-5). Sixty-one components were characterized comprising 78.7% and 76.2% of the oil composition, respectively. The major oxygenated sesquiterpenes identified possessed structures based on valerenic acid skeleton, valerenal being the most representative. The oil composition significantly varied due to drying procedure. Dried roots contained high amount of free isovaleric acid (13.0%) in comparison to fresh roots (0.2%) which may be due to enzymatic hydrolysis of isovaleric esters upon drying. Higher contents of borneol and sesquiterpene hydrocarbons were also found in dried root oil. Dried roots were also steam distilled and three oil fractions were collected in equal time periods during the steam distillation. It was possible to mod...