Differentiation of Belamcandae Rhizoma and Iridis Tectori Rhizoma by Thin-Layer Chromatography and High-Performance Liquid Chromatography

Objective:Belamcandae Rhizoma and Iridis Tectori Rhizoma are easily confused with each other.The main objective of this study is to distinguish them using chemical analysis.Materials and Methods:Thin-layer chromatography(TLC)and high-performance liquid chromatography(HPLC)fingerprint methods were established to compare the chemical profile,while HPLC quantitation was used to determine the contents of three isoflavones in thirty batches of Belamcandae Rhizoma and Iridis Tectori Rhizoma samples.Results:The two herbs could be distinguished by TLC using acetic acid-n hexane-ethyl acetate(1:90:80 v/v/v)as the mobile phase,according to the fluorescent band under 366 nm at R_(f) 0.2.In total,12 compounds were identified in the 24-min HPLC fingerprint.The similarity coefficient between the two herbs was 0.54±0.01.Mangiferin(1),tectoridin(2),iridin(3),irigenin(5),irisflorentin(6),and iristectorin A(9)were the main peaks in Belamcandae Rhizoma,while tectoridin(2)and tectorigenin(4)were the major peaks in Iridis Tectori Rhizoma.The contents of 2 in Iridis Tectori Rhizoma(2.50±0.20%)were 8.93 times higher than that of Belamcandae Rhizoma(0.28±0.08%),while the ones of 5 and 6 were slightly lower in Iridis Tectori Rhizoma.Conclusions:The study established fast and effective methods to distinguish Belamcandae Rhizoma from Iridis Tectori Rhizoma.

Anti-Acinetobacter baumannii activity of Rumex crispus L. and Rumex sanguineus L. extracts

Objective:To examine the effect of Rumex crispus(R.crispus)and Rumex sanguineus(R.sanguineus)plant extracts against isolates of Acinetobacter baumannii(A.baumannii)from wounds,including multidrug-resistant strains.Methods:Six prepared Rumex extracts were subjected to liquid chromatography-tandem mass spectrometry.Antimicrobial activity of extracts and pure compounds(catechin,quercetin,isoquercitrin,emodin,and gallic acid)was examined by a microtiter plate method,while for determination of compound binary combinations activity a checkerboard method was applied.Active fractions of extracts were detected by agar-overlay high-performance thinlayer chromatography-bioautography assay followed by liquid chromatography-diode array detection-mass spectrometry analysis.Results:A total of 28 compounds were detected in two extracts of R.crispus and 26 compounds in four different R.sanguineus extracts,with catechin as a dominant component.Anti-A.baumannii activity was confirmed for all six R.sanguineus and R.crispus extracts at the concentration range from 1 to 4 mg/mL.Neither examined single compounds nor their binary combinations exhibited an anti-A.baumannii activity(MIC>256μg/mL).The bioautography showed that fractions with the most prominent anti-A.baumannii activity tended to contain more polar compounds,predominantly flavonol(quercetin and kaempherol)glycosides;but also fractions containing flavanone(eriodictyol)glycosides and anthraquinone(emodin)glycosides;and less polar eriodictyol aglycone.Conclusions:The results justify and elucidate the traditional application of R.sanguineus and R.crispus extracts for wound healing,indicating the necessity for their further examination in combat against multidrug-resistant A.baumannii isolates from wounds.

Qualitative Identification and Content Determination of Psoralen in Ficus Pandurata Hance

[Objectives]This study aimed to establish a qualitative identification and content determination method for psoralen in Ficus pandurata Hance and compare the psoralen contents in roots,stems and leaves of F.pandurate Hance and Ficus pandurata Hance var.holophylla Migo.[Methods]Thin-layer chromatography(TLC)was used for qualitative identification,and the content of psoralen was determined by high-performance liquid chromatography.The chromatographic conditions were as follows:column,Agilent ZORBAX Eclipse Plus C18(250 mm×4.6 mm,5μm);mobile phase,methanol-water(55∶45);flow rate,1 mL/min;detection wavelength,246 nm;column temperature,30℃;and injection volume,10μL.[Results]The TLC chromatogram of F.pandurate Hance showed clear spots and good separation.The concentration of psoralen detected in the range of 2.06-41.20μg/mL had a good linear relationship with the peak area(r=0.9999).The RSD values of the precision,stability and reproducibility tests were all less than 2%.The average recovery rate was 100.2%(RSD=1.13%,n=6).[Conclusions]The established method is simple,easy,accurate and reproducible.It can be used for quality control of F.pandurate Hance.

Antimicrobial activity of Cannabis sativa,Thuja orientalis and Psidium guajava leaf extracts against methicillin-resistant Staphylococcus aureus

Objective： This study examined the antimicrobial activity of Cannabis sativa, Thuja orientalis and Psidium guajava against methicillin-resistant Staphylococcus aureus （MRSA） and used a standardized purification protocol to determine the presence and abundance of bioactive compounds in the leaf extracts. Methods： In vitro antimicrobial activities of the ethanolic extracts of C sativa, T. orientalis and P. guajava were tested against MRSA. The presence of bioactive molecules in these three leaves was evaluated using biochemical assays and high-performance thin-layer chromatography （HPTLC）. Results： Resistance to methicillin, penicillin, oxacillin and cefoxitin was observed in each of the clinical and nonclinical MRSA isolates. However, they were still vulnerable to vancomydn. Used individually, the 50% extract of each plant leaf inhibited MRSA growth. A profound synergism was observed when C sativa was used in combination with T. orientalis （1：1 ） and when P. guajava was used in combination with T. orientalis （1：1 ）. This was shown by larger zones of inhibition. This synergism was probably due to the combined inhibitory effect of phenolics present in the leaf extracts （i.e., quercetin and gallic acid） and catechin, as detected by HPTLC. Conclusion： The leaf extracts of C sativa, T. orientalis and P. guajava had potential for the control of both hospital- and community-acquired MRSA. Moreover, the inhibitory effect was enhanced when extracts were used in combination.

Marker-based standardization and investigation of nutraceutical potential of Indian propolis

OBJECTIVE： Propolis, a resinous material collected by honey bees from various plants, has been explored globally for its medicinal and nutritional properties. However, research over Indian propolis is at infancy. This study was designed to investigate nutraceutical potential of Indian propolis. METHODS： In the present study, propolis extract was standardized with respect to markers caffeic acid phenethyl ester, caffeic acid, galangin, luteolin, curcumin, apigenin, pinocembrin and quercetin by new high-performance thin-layer chromatographic （HPTLC） methods. The physico-chemical analysis, residues analysis and in vitro antioxidant activity analysis were performed. Nutraceutical value was examined in terms of fats, fibers, minerals, proteins, polysaccharides, total carbohydrates, and energy value. RESULTS： The developed HPTLC methods were found to be simple, reliable accurate, and the validation parameters were within the limits of the International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use guidelines. Macerated ethanolic extract of propolis （MEEP） was found to have polyphenolic content of （20.99 ＋ 0.24） mg/g and flavonoids content of （8.39 ＋ 0.04） mg/g. MEEP was found to comprise of （283.33 ＋ 51.31） g/kg fats, （30.07 ＋ 7.30） g/kg fibers, （102.56 ＋ 2.84） g/kg proteins and （389.36 ＋ 57.50） g/kg carbohydrate with a calorie value of （38 409.33 ＋ 6 169.80） kJ/kg. It was found that Indian propolis exhibited high nutraceutical value and showed absence of pesticides and heavy metals. The MEEP showed in vitro antioxidant activity with inhibitory concentration of （12.24 ＋ 4.64） pg/mL. CONCLUSION： The present work explores Indian propolis as a potential nutritious candidate. The proposed analytical methods can be applied in future screening of the quality of Indian propolis.

Development of Diagnostic Microscopic and Chemical Markers of Some Euphorbia Latexes

The latexes of the three Euphorbia species, namely E. antiquorum L., E. nerifolia L., and E. tirucalli L., are highly valued in the Indian system of medicine as purgatives, in addition to their specific and distinct therapeutic activities. In order to distinguish these latexes and develop their diagnostic microscopic and chemical markers, we performed extensive chemical and microscopic studies. The three latexes differ significantly in their microscopic features by exhibiting characteristic starch grain patterns. Although amoebic structures were found to be characteristic of E. antiquorum, dumb-bell and oval structures are characteristic of E. nerifolia and E. tirucalli, respectively. In addition, these latexes showed bone-shaped structures as a common feature, but these differed considerably in their length （10-60, 30-55, and 50-70 μm in length in E. antiquorum, E. nerifolia, and E. tirucalli, respectively）. The chemical markers nerifoliene and euphol were found to be common to both E. antiquorum and E. nerifolia, whereas euphol is the only marker for E. tirucalli. A reverse-phase high-performance thin-layer chromatographic （HPTLC） method was developed to distinguish these three latexes and to generate their standard fingerprinting patterns. Most significantly, the markers nerifoliene and euphol could be resolved by RP-18 F254s precoated aluminium plates and the latexes have been quantitatively estimated with respect to these markers. The developed microscopic, chemical and HPTLC patterns can be used to distinguish the three latexes.

Chenopodium ambrosioides induces an endothelium-dependent relaxation of rat isolated aorta

Objective: This study aims to evaluate the vasodilatory effect of Chenopodium ambrosioides on the isolated rat aorta, and to explore its mechanism of action.Methods: The vasorelaxant effect and the mode of action of various extracts from the leaves of C. ambrosioides were evaluated on thoracic aortic rings isolated from Wistar rats. In addition, ethyl acetate and methanol fractions were analyzed, using thin-layer chromatography and high-performance liquid chromatography techniques, for their polyphenolic content.Results: The various active extracts of C. ambrosioides at four concentrations(10^(-3), 10^(-2), 10^(-1) and 1 mg/mL) relaxed the contraction elicited by phenylephrine, in a concentration-dependent manner.This effect seems to be endothelium-dependent, since the vasodilatory effect was entirely absent in denuded aortic rings. The vasorelaxant effect of the methanol fraction(MF) of C. ambrosioides at 1 mg/mL was also inhibited by atropine and tetraethylammonium. This effect remained unchanged by Nx-nitro-L-arginine methyl ester hydrochloride and glibenclamide. The preliminary phytochemical analysis showed that the leaves of C. ambrosioides are rich in phenolic and flavonoid derivatives.Conclusion: These results suggest that the MF of C. ambrosioides produces an endothelium-dependent relaxation of the isolated rat aorta, which is thought to be mediated mainly through stimulation of the muscarinic receptors, and probably involving the opening of Ca^(2+)-activated potassium channels.