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The cox2 locus of the primitive angiosperm plant Acorus calamus: molecular structure, transcript processing and RNA editing.

Acorus calamus, or sweet flag, is a semiaquatic plant of uncertain taxonomic position. Molecular phylogenetic analysis using plastid rbcL sequences have suggested that Acorus calamus might be the most ancient surviving representative of the ancestral monocotyledonous plants. In order to provide molecular and phylogenetic data for the mitochondrial genetic system of Acorus, we have determined the structure of a mitochondrial locus, the cytochrome oxidase subunit II gene cox2. The Acorus cox2 gene harbors an unusually small group II intron, the smallest plant mitochondrial intron known to date. The transcript undergoes C-to-U RNA editing at eight sites. One of these sites is likely to play a dual functional role in both intron splicing and protein function. The 3' end of the mature transcript folds into a characteristic stem-loop structure that is presumably required for mitochondrial mRNA stability. Phylogenetic analysis of the cox2 sequence data, as well as the unusual intron structure, all support an evolutionarily isolated position for Acorus calamus.

Base Sequence↗

Novel lectins from rhizomes of two Acorus species with mitogenic activity and inhibitory potential towards murine cancer cell lines.

Two novel lectins were purified from rhizomes of two sweet flag species, namely Acorus calamus (Linn.) and Acorus gramineus (Solandin Ait.) by affinity chromatography on mannose linked epoxy-activated Sepharose 6B. The apparent molecular mass of the lectins, as determined by gel filtration chromatography, was 56 kDa for ACL and 55 kDa for AGL. In SDS-PAGE, pH 8.3, both lectins migrated with a subunit molecular mass of 13.6 kDa and 13.5 kDa, respectively, under reducing and non-reducing conditions thus indicating the absence of disulphide linkages. Acorus lectins readily agglutinated rabbit, rat and guinea pig erythrocytes. Both ACL and AGL also reacted with RBCs from sheep, goat and human ABO blood groups after neuraminidase treatment. ACL and AGL were inhibited by mannose/glucose and their derivatives. The most effective inhibitor was methyl-alpha-D-mannopyranoside. Acorus lectins were stable up to 55 degrees C, did not require metal ions for their activity and were also affected by high concentrations of denaturants like urea, thiourea and guanidine-HCl. These lectins showed potent mitogenic activity towards mouse splenocytes and human lymphocytes. Both ACL and AGL also significantly inhibited the growth of J774, a murine macrophage cancer cell-line and to lesser extent WEHI-279, a B-cell lymphoma.

Acorus↗

Developmental anatomy of the root cortex of the basal monocotyledon, Acorus calamus (Acorales, Acoraceae).

BACKGROUND AND AIMS: The anatomical structure and development of adventitious roots were analysed in the basal monocotyledon, Acorus calamus, to determine to what extent those features are related to phylogenetic position. METHODS: Root specimens were harvested and sectioned, either with a hand microtome or freehand, at varying distances from the root tip and examined under the microscope using a variety of staining techniques. KEY RESULTS: Roots of Acorus calamus possess a unique set of developmental characteristics that produce some traits similar to those of another basal angiosperm group, Nymphaeales. The root apical meristem organization seems to be intermediate between that of a closed and an open monocotyledonous root apical meristem organization. The open-type root apical meristem consists of a curved zone of cortical initials and epidermal initials overlying the vascular cylinder initials; the epidermal part of the meristem varies in its association with the cortical initials and columellar initials of the promeristem. The cortex develops an endodermis with only Casparian bands, a dimorphic exodermis with Casparian bands and suberin lamellae, and a polygonal aerenchyma by differential expansion, as also observed in the Nymphaeales and some dicotyledonous species. The stele has characteristics like those of members of the Nymphaeaceae. CONCLUSIONS: Specific anatomical and developmental attributes of Acorus roots seem to be related to the phylogenetic position of this genus.

Acorus↗

[Microscopic identification of medicinal species of Acorus].

A study on the identification of commercial drugs of medicinal species of Acorus collected from 10 different areas had been carried out. The result showed that the commercial drugs were mainly derived from three species, i. e., Acorus tatarinowii, A. calamus L. and A. macrospadicesu (Yamamoto) F. N. Wei et Y. K. Li. With the digital imaging technique, the paper visually reflected the microscopic structures of medicinal plants of Acorus, and can be used for identifying the market drugs.

Acorus↗

Protective effect of acorus calamus against acrylamide induced neurotoxicity.

Exposure of rats to acrylamide (ACR) caused hind limb paralysis in 58% of the animals on day 10 and decreased behavioural parameters, namely distance travelled, ambulatory time, stereotypic time and basal stereotypic movements compared with the control group. These rats also had a decrease in the reduced glutathione (GSH) content and glutathione-S-transferase (GST) activity in the corpus striatum and an increase in striatal dopamine receptors, as evident by an increase in the binding of 3H-spiperone to striatal membranes. Treatment with the ethanol:water (1:1) extract of the rhizomes of Acorus calamus (AC-002) increased the GSH content and GST activity in the corpus striatum while insignificant changes were observed in other parameters. Rats treated with ACR and AC-002 in combination had a lower incidence of paralysis (18%) compared with those treated with ACR alone on day 10 of the experiment. The rats also showed a partial recovery in other behavioural parameters. The levels of GSH content and GST activity increased in the corpus striatum, while the dopamine receptors decreased compared with the ACR treated rats. The results suggest that the neurobehavioural changes produced by ACR may be prevented following treatment with Acorus calamus rhizomes.

Acorus↗

Rapid determination of essential oil in Acorus tatarinowii Schott. by pressurized hot water extraction followed by solid-phase microextraction and gas chromatography-mass spectrometry.

A simple, rapid, efficient and inexpensive method for the determination of essential oil in a traditional Chinese medicine (TCM), Acorus Tatarinowii Schott. was developed by using pressurized hot water extraction (PHWE) combined with solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS). Acorus Tatarinowii Schott. (0.050 g) was subjected to dynamic extraction with water at 50 bar, 150 degrees C and 1.0 ml/min for 5.0 min, and then the essential oil in the aqueous extract were extracted by SPME fibers at 60 degrees C for 10 min, finally the compounds on the fiber were desorbed and analyzed by GC-MS. The PHWE and SPME parameters were studied. The key active compound of alpha-asarone in the TCM samples from three different growing areas was quantitatively analyzed by external standard method. Compared to steam distillation (SD), the proposed method required little time (only 15 min) to prepare sample. Moreover, little sample mass and no organic solvent was needed. The present method provided good repeatability (R.S.D. less than 13.0%) and recovery (92% for alpha-asarone). It has been shown that PHWE-SPME-GC-MS is an alternative method for determination of essential oils in TCMs and a potential tool for TCM quality assessment.

Acorus↗

Chemical composition of the essential oil and supercritical CO2 extract of Commiphora myrrha (Nees) Engl. and of Acorus calamus L.

Volatile concentrates from the oleo-gum resin of Commiphora myrrha (Nees) Engl. and from the rhizomes of Acorus calamus were isolated by supercritical extraction with carbon dioxide. The volatile oil of myrrh was obtained at 9.0 MPa and 50 degrees C and at a CO2 flow of 1.5 kg/h. Acorus calamus was extracted at 9.0 MPa and 45 degrees C and at a CO2 flow of 1.6 kg/h. In both cases, an oil devoid of cuticular waxes was obtained with a single depressurization stage. The SFE myrrh oil had a yield, Y, of 3.2%. Its main components, identified and quantified by GC/MS, were furanoeudesma-1,3-diene, 34.9%; lindestrene, 12.9%; curzerene, 8.5%; and germacrone, 5.8%. The essential oils from the same starting material by hydrodistillation, HD, (Y = 2.8%) and by steam distillation, SD, (Y = 0.4%) were quite similar to the SFE extract. The main components of the SFE oil of A. calamus (Y = 3.5%) were acorenone, 13.4%; iso-acorone, 11.6%; (Z)-sesquilavandulol, 11.0%; dehydroxy isocalamendiol, 7.7%; and beta-asarone, 5.5%. The comparison with hydrodistilled (Y = 1.8%) and steam distilled (Y = 1.0%) oils revealed large differences in the content of iso-acorone and crypto-acorone.

Acorus↗

Study of antiepileptic effect of extracts from Acorus tatarinowii Schott.

PURPOSE: To study the antiepileptic properties of extracts from rhizomes of Acorus tatarinowii Schott (ATS). METHODS: The decoction and volatile oil were extracted from rhizomes of ATS by traditional decocting and supercritical CO(2) fluid extraction (SFE-CO(2)) methods. Maximal electroshock (MES), pentylenetetrazol (PTZ) maximal seizure, and prolonged PTZ kindling models were used to test their anticonvulsive properties. The gamma-aminobutyric acid (GABA) immunohistochemical reaction (IR) was used to study GABAergic neuron changes in the PTZ kindling model and the effects of treatment. RESULTS: Both decoction (dose; 10-20 g/kg) and volatile oil (1.25 g/kg) of ATS decreased the convulsive rate significantly in the MES model. Decoction of ATS was shown to be effective in the PTZ model with both decreased convulsive and mortality rates. The volatile oil of ATS failed to prevent seizures in the dose range tested, although prolonged seizure latency and decreased mortality were found at a dose of 1.25 g/kg. In the PTZ kindling model, GABA-IR neurons decreased obviously compared with the normal group. In the groups treated with the decoction and volatile oil, the seizure intensity decreased significantly after treatment. Increased GABA-IR neurons also were found when compared with PTZ kindling controls. Morphologic observation also showed that GABA-IR neuron damage was less severe in the drug-treated groups. CONCLUSIONS: Both decoction and volatile oil extracted from the rhizome of Acorus tatarinowii Schott have anticonvulsive effects. The volatile oil is shown to be less effective for PTZ-induced convulsions. Both extracts can prevent convulsions as well as convulsion-related GABAergic neuron damage in the brain in the prolonged PTZ kindling model.

Acorus↗

Protective effect of Acorus calamus LINN on free radical scavengers and lipid peroxidation in discrete regions of brain against noise stress exposed rat.

Exposure to continuous loud noise is a serious health problem due to excess production of oxygen free radicals. In medical research, more attention is paid to the antioxidant properties of medicinal plants to minimize the harmful effects of radicals. The aim of this study was to evaluate the protective effect of both ethyl acetate and methanolic extract of Acorus calamus LINN against noise stress (30 d, 100 dBA/4h/d) induced changes in the rat brain. We measured the activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and the levels of reduced glutathione (GSH), vitamin C, vitamin E, protein thiols and lipid peroxidation (LPO) for the evaluation of oxidative stress status in discrete regions of the rat brain like cerebral cortex, cerebellum, pons-medulla, midbrain, hippocampus and hypothalamus. The results indicated that during exposure of noisy environment ROS generation led to increase in corticosterone, LPO and SOD, but decrease in CAT, GPx, GSH, protein thiols, vitamins C and E levels. Both the ethyl acetate and methanolic extract of Acorus calamus protected most of the changes in the rat brain induced by noise-stress.

Acetates↗

[Effect of Acorus tatarinowii Schott. on gastrointestinal myoelectric activity in rats].

The effect of Acorus tatarinowii on the gastrointestinal myoelectric activity and mechanism in rats was studied electrio-physiologically. The result indicates that Acorus tatarinowii helps inhibit the gastrointestinal myoelectric activity. This inhibitory effect is believed to take place through the blocking of cholinergic M and non-cholinergic M receptors, and has nothing to do with adrenergic alpha and beta receptors.

Acorus↗

[Study on GC-MS fingerprint analysis in rhizome of volatile oil of Acorus tatarinowii].

OBJECTIVE: To establish the method of fingerprint analysis on volatile oil in rhizome of Acorus tatarinowii by GC-MS, and to study the main characteristic components. METHOD: The main components of 10 samples were determined by GC-MS. RESULT: The injector temperature was 250 degrees C. The interface temperature was 230 degrees C. The column flow was 1.3 mL x min(-1). The column pressure was 80 kPa. The detector volt was 1.4 kV. The temperature rate was 3 degrees C x min(-1). And the main characteristic components were composed of the methyleugenol (2.13%), cis-methylisoeugenol (4.48%), trans-methylisoeugenol (0.82%), gamma-asarone (4.51%), beta-asarone (66.15%), alpha-asarone (6.35%). And the RSD of precision and reproducibility and stability was almost in the range of 5%. CONCLUSION: The method is reliable, accurate and can be used for fingerprint analysis of volatile oil of Acorus tatarinowii.

Acorus↗

Screening and isolation of antibiotic resistance inhibitors from herb materials IV-resistance inhibitors from Anetheum graveolens and Acorus gramineus.

The hexane fractions from methanolic extracts of Anetheum graveolens L. (Umbelliferae) and Acorus gramineus Soland. (Araceae), revealed potent inhibitory activities against the resistance of multi-drug resistant Staphylococcus aureus SA2 when combined with ampicillin (Am) or chloramphenicol (Cm). As active principles, carvone and the liquid mixture containing carvone from Anetheum graveolens L. and a liquid mixture mainly consisting of benzoic acid phenylmethyl ester (benzyl benzoate) from Acorus gramineus Soland, were identified. They showed resistance inhibition at the level of 20-50 micrograms/ml when combined with 100 or 50 micrograms/ml of Am or Cm, respectively.

Ampicillin↗

Phylogenetic analysis of rbcL sequences identifies Acorus calamus as the primal extant monocotyledon.

The identity of the oldest lineage of monocotyledons is a subject of debate. Alternative interpretations of morphological homologies are variously consistent with proposals that species of Alismatanae, Dioscoreales, or Melanthiales were the earliest descendants of the first monocotyledons. We present phylogenetic analyses based on DNA sequences of the plastid locus rbcL in which Acorus calamus, an herb with unspecialized floral features and of uncertain affinities, is supported as a member of the oldest extant lineage of monocotyledons. This conclusion is consistent with a substantial body of morphological, anatomical, and embryological evidence and offers an explanation for the failure to identify any close relationship between Acorus and other genera.

Chloroplasts↗

DNA profiling of Acorus calamus chemotypes differing in essential oil composition.

The phylogenetic relationship of Acorus gramineus and three types of Acorus calamus was analyzed by comparing the 700 bp sequence of a 5S-rRNA gene spacer region. Although there was no intra-specific variation in the essential oil profile of A. gramineus which contained a phenylpropanoid (Z-asarone) as a predominant constituent, A. calamus was classified into two chemotypes: chemotype A in which Z-asarone is a major essential oil constituent and chemotype B which contained sesquiterpenoids predominantly. An intermediate type (M) of these two chemotypes in various ratios was also observable. The NJ tree constructed based on the sequences revealed that A. gramineus was clearly distinguished from any of the chemotypes of A. calamus and that the phylogenetic relationship predicted by the spacer region data correlated well with the essential oil chemotype pattern of A. calamus.

Base Sequence↗

Rapid assessment of beta-asarone content of Acorus calamus by micellar electrokinetic capillary chromatography.

This report outlines a rapid, reproducible method for the determination of beta-asarone, a known carcinogen, using micellar electrokinetic capillary chromatography (MEKC)-UV-vis absorbance and a simple alcohol extraction. The MEKC method is based on a running buffer comprised of 100 mM sodium dodecyl sulfate (SDS), pH 10. The method is reproducible and provides baseline separation of alpha-asarone and beta-asarone. This protocol was used to determine the beta-asarone content of Acorus calamus rhizome of a diploid variety harvested from the wetlands of the United States and the triploid variety from India obtained commercially. The results indicate raw product that originated from India contained 4.4% w/w beta-asarone, while that from the United States contained 0.2% w/w beta-asarone. Neither sample contained detectable concentrations of alpha-asarone. This is the first report of the use of MEKC to determine asarone in a natural source.

Acorus↗

Antispasmodic effect of Acorus calamus Linn. is mediated through calcium channel blockade.

Acorus calamus Linn. (Araceae) is a native of Central Asia and Eastern Europe and has widespread use in the traditional system of medicine for gastrointestinal disorders such as colic pain and diarrhoea. This study was aimed at providing a possible pharmacological basis to the use of this plant as an antispasmodic and antidiarrhoeal. In the isolated rabbit jejunum preparation the crude extract (Ac.Cr), which tested positive for the presence of alkaloid, saponins and tannins, caused inhibition of spontaneous and high K(+) (80 mm)-induced contractions, with respective EC(50) values of 0.42 +/- 0.06 and 0.13 +/- 0.04 mg/mL (mean +/- SEM; n = 6-8), thus showing spasmolytic activity, mediated possibly through calcium channel blockade (CCB). The CCB activity was confirmed when pre-treatment of the tissue with Ac.Cr (0.3-1.0 mg/mL) caused a rightward shift in the Ca(++) dose-response curves similar to that caused by verapamil, a standard calcium channel blocker. Activity-directed fractionation revealed that the CCB activity was concentrated in the n-hexane fraction while the ethylacetate fraction was less potent. These results suggest that the spasmolytic effect of the plant extract is mediated through the presence of CCB-like constituent(s) which is concentrated in the n-hexane fraction and this study provides a strong mechanistic base for its traditional use in gastrointestinal disorders such as colic pain and diarrhoea.

Acorus↗

Fungicidal property of active component derived from Acorus gramineus rhizome against phytopathogenic fungi.

The fungicidal property of Acorus gramineus rhizome-derived materials against Botrytis cineria, Erysiphe graminis, Phytophthora infestans, Puccinia recondita, Pyricularia grisea, and Rhizoctonia solani was tested using a whole plant method in vivo, and was compared with a synthetic fungicide and seven commercially available A. gramineus rhizome-derived components. The responses varied according to the plant pathogen tested. At 2000 mg/l, the hexane fraction of A. gramineus rhizomes showed strong fungicidal activities against R. solani and P. infestans. The results indicated at least one of the fungicidal actions of alpha-asarone and asaronaldehyde derived from A. gramineus rhizomes.

Acorus↗

Identification of an EcoRI restriction site for a rapid and precise determination of beta-asarone-free Acorus calamus cytotypes.

Calamus (Acorus calamus L., Araceae) is an aromatic herb, indigenous to Central Asia and Eastern Europe. The fragrant oils obtained by alcoholic extraction of the rhizome are mainly used in the pharmaceutical and oenological industries. Nevertheless, the occurrence of beta-asarone [(Z)-1,2,4-trimethoxy-5-prop-1-enyl-benzene] limits the possibility of its use due to the carcinogenic properties of this compound. The aim of this work was to identify a diploid beta-asarone-free A. calamus by using chemical and molecular approaches. For these purposes alcoholic extracts of both diploid and triploid A. calamus were analyzed by gas chromatography-mass spectrometry (GC-MS) and comparison of the 700 bp sequence of the non-transcribed spacer (NTS) in the 5S-rRNA gene was also performed. Alcoholic extracts of the triploid A. calamus were characterized by a higher percentage of beta-asarone (11%), which was the main compound, followed by higher percentages of camphene (2.27%), E-beta-ocimene (3.28%), camphor (1.54%), calarene (1.42%), alpha-selinene (5.02%) and tau-cadinol (2.00%), when compared to the diploid A. calamus. The latter had higher percentages of iso-shyobunone (8.62%), beta-sesquiphellandrene (3.28%), preiso calamendiol (22.81%) and acorone (26.33%), and completely lacked of beta-asarone. The 5S-rRNA spacer region of both diploid and triploid A. calamus were amplified by PCR using a pair of primers located at the 3' and 5' ends of the coding sequence of 5S-rRNA gene. The resulting PCR products (about 700 bp) were gel purified, subcloned into pGEM-T Easy vector and sequenced. By aligning the isolated nucleotide sequences of the two varieties and the sequences from different A. calamus chemotypes present in Genbank, sequence diversities were found in the spacer region. Furthermore, the PCR products were digested by using EcoRI. The restriction profile of the spacer domain resulted different for the two cytotypes. Along with chemical analysis of alcoholic extracts, sequence analysis coupled to restriction mapping was demonstrated to represent a powerful tool to distinguish the A. calamus diploid cytotype from the others. The security and effective usage of the diploid beta-asarone-free A. calamus was also discussed.

Acorus↗