Preparative Layer Chromatography Research Articles

GC/MS Analysis and Isolation of Stigmasterol from Viola Odorata Cultivated in Iraq

Viola odorata is a viola family flowering plant native to Europe and Asia. This is a small, hardy, herbaceous, perennial plant. It is also known as sweet violet and English violet. Numerous pharmacological properties of this flowering plant include anticancer, antibacterial, anti-inflammatory, antioxidant and antipyretic properties. Previous literature review of different parts of Viola odorata resulted in detection of different chemical constituents such as flavonoids (Quercetin, kaempferol), glycosides (Rutin) and terpenoids (stigma sterol). Stigmasterol was detected using TLC and GC-MS in this study. It was isolated from Viola odorata using preparative layer chromatography. HPLC and FTIR analysis confirmed the isolated Stigmasterol

Isolation, Identification, and Quantification of Two Compounds from Cassia glauca Cultivated in Iraq

The Cassia glauca Lam. is the tree that belongs to the Fabaceae family and is native to India has many uses in indigenous systems of medicine, folk medicine, and traditional Brazilian medicine. Has many pharmacological activities such as anti-diabetic, antibacterial, antifungal, antioxidant, anti-hemolytic, anticancer, cardio-protective, and Hepato-protection. The aim of study is to Isolation, identification, and quantification of some compounds from aerial parts of Cassia glauca since no phytochemical investigation had previously been done in Iraq for this plant. The aerial parts were defatted in n. hexane for 48 hours. The defatted materials were extracted in 85% ethanol using the hot method (soxhlet), then the extract was fractionated using different solvents (chloroform, ethyl acetate, and n-butanol). High-performance liquid chromatography (HPLC), was used for identification and quantification by using authenticated standards, and preparative layer chromatography (PLC) was used for the isolation of the identified compounds. The isolated compounds were identified after isolation by liquid chromatography Mass Spectrometry LC-MS/MS-Q-TOF method. The different chromatographic and spectroscopic methods results indicate the presence of luteolin and chlorogenic acid in the ethyl acetate fraction and n-butanol fraction respectively and estimated the quantity as (130.77µg/1g) for luteolin, and (0.0006%) for chlorogenic acid from 50g of plant material.

Antiangiogenic activity and the isolation of five phenolic compounds from Euphorbia milii ethyl acetate solvent extract

Background: Euphorbia milii a medicinal plant from Euphorbia genus, has been used traditionally as a cure for cancer, warts and as a hepatoprotective agent in tropical areas like China, Myanmar, Brazil, and Pakistan. Previous literature investigated the effect of E. milii flower extract in some types of cancer cell lines. Objective: This study aims to evaluate the antiangiogenic activity of the ethyl acetate fraction of the methanolic crude extract of E. milii, in addition to the detection and isolation of some phenolic compounds. Materials and Methods: Detection of the phenolic compounds was achieved using TLC and HPLC chromatographic methods. Isolation was achieved by preparative layer chromatography based on Rf values. The isolated compounds were identified through HPLC and TLC, in addition to FTIR. The anti-angiogenic activity was evaluated employing rat aorta ring anti-angiogenesis assay. Results: Five phenolic compounds were isolated in this study namely, Luteolin, Apigenin, Isorhamnetin, Coumaric acid and Quercitrin. However phenolic compounds detected via TLC were, Silybin, Luteolin, Apigenin, Isorhamnetin, Coumaric acid and Quercitrin. Interestingly, HPLC detected Coumarin, Rutin, Catechin and Quercetin. Anti-angiogenesis assay found that E. milii ethyl acetate fraction inhibited new blood vessel growth significantly (p<0.05) by 94 + 3.6% as compared with DMSO negative control (0%), while when compared with Suramin positive control (100%) it showed no significant difference (p>0.05). Conclusion: E. milii showed significant anti angiogenic activity of its ethyl acetate fraction, suggesting the association of E. milii traditional use as anticancer and phenolic content, current data must be further evaluated.

Determination and Isolation of Valuable Bioactive Compound (lupeol) from Portulacaria afra Jacq.

Portulacaria afra is a small succulent tree, previously belonging to the Portulacaceae family, but with further studies, the plant transferred to the Didieracea family. P. afra was used as an ornamental, vegetable, and ethnomedicinal plant. Uses of the plant by rural South Africans to treat chronic skin conditions and rashes, alleviate exhaustion, and aid in treating TB and diarrhea have been documented in folklore. According to pharmaceutical research, plant extracts off er a wide range of remedial outcomes, such as antidiabetic, antifungal, antibacterial, anticancer, antioxidant, and anti-infl ammatory. The study aims to determine some bioactive constituents responsible for pharmacological activities and traditional usefulness. Thinlayer chromatography (TLC) is used for detecting lupeol by specifi c reagents; a p-anisaldehyde sulfuric acid reagent and 10% methanolic sulfuric acid. And high-performance liquid chromatography was used to detect pentacyclic triterpenoids (lupeol) in the n-hexane. The lupeol was isolated by preparative layer chromatography (PLC). Testing the effi cacy of the separation method, the isolated compounds have been identifi ed and characterized by diff erent chromatographic and chemical analyses (TLC, ATR-FTIR, LC-CMS-APCI+, and 1H-NMR).

Halorotetin A: A Novel Terpenoid Compound Isolated from Ascidian Halocynthia rotetzi Exhibits the Inhibition Activity on Tumor Cell Proliferation.

Halocynthia roretzi, the edible ascidian, has been demonstrated to be an important source of bioactive natural metabolites. Here, we reported a novel terpenoid compound named Halorotetin A that was isolated from tunic ethanol extract of H. roretzi by silica gel column chromatography, preparative layer chromatography (PLC), and semipreparative-HPLC. 1H and 13C NMRs, 1H-1H COSY, HSQC, HMBC, NOESY, and HRESIMS profiles revealed that Halorotetin A was a novel terpenoid compound with antitumor potentials. We therefore treated the culture cells with Halorotetin A and found that it significantly inhibited the proliferation of a series of tumor cells by exerting cytotoxicity, especially for the liver carcinoma cell line (HepG-2 cells). Further studies revealed that Halorotetin A affected the expression of several genes associated with the development of hepatocellular carcinoma (HCC), including oncogenes (c-myc and c-met) and HCC suppressor genes (TP53 and KEAP1). In addition, we compared the cytotoxicities of Halorotetin A and doxorubicin on HepG-2 cells. To our surprise, the cytotoxicities of Halorotetin A and doxorubicin on HepG-2 cells were similar at the same concentration and Halorotetin A did not significantly reduce the viability of the normal cells. Thus, our study identified a novel compound that significantly inhibited the proliferation of tumor cells, which provided the basis for the discovery of leading compounds for antitumor drugs.

Isolation of beta-sitosterol and evaluation of antioxidant Activity of Iraqi Campsis grandiflora flowers.

Campsis grandiflora (Bignoniaceae) is a fast growing deciduous climber, the dried flowers have been used as a carminative, blood tonic, and febrifuge in Chinese traditional medicine. This plant has an anti-inflammatory, anti-oxidant, anti-depressant, and anti-bacterial effect; with a beneficial role in stagnant blood and endometriosis conditions. In this study, the detection of beta-sitosterol in the hexane extract of Iraqi C.grandiflora flowers was performed using thin layer chromatography (TLC) and high performance liquid chromatography(HPLC); while the isolation done by preparative layer chromatography then structure elucidation of isolated compound was done by FTIR and 1HNMR. Furthermore, assessment of the anti-oxidant activity of the ethyl acetate extract of Iraqi C.grandiflora flowers using three different methods and total flavonoid content, then measuring the pearson’s correlation coefficient between these methods. The results showed that the hexane extract of Iraqi C.grandiflora flowers contain beta-sitosterol compound and the ethyl acetate extract of this plant possesses an excellent anti-oxidant effect using the single-electron transfer (SET) pathway in scavenging the free radicals, and this activity attributed to the potent antioxidant i.e. polyphenols.

Synthesis and cytotoxicity of the boron carrier pentagamaboronon-0-ol for boron neutron capture therapy against breast cancer.

Boronic acid-containing curcumin analog, pentagamaboronon-0 (PGB-0), acts as a potential boron-carrier agent but has limited water solubility. Thus, a new compound (PGB-0-ol) with better chemical and pharmacological properties than PGB-0 has been synthesized. Molecular docking was performed using a molecular operating environment. Prediction of PGB-0-ol absorption, distribution, metabolism, and excretion (ADME) was performed using pkCSM software. PGB-0-ol was synthesized by adding NaBH4 to PGB-0 and stirring for 1 h. The crude PGB-0-ol was purified using preparative layer chromatography. Cell viability was evaluated using the trypan blue exclusion assay. In comparison to PGB-0 based on molecular docking study, PGB-0-ol could interact in with several cancer biomarkers, such as human epidermal growth factor2 epidermal growth factor receptor, IκB kinase, folate receptor-α, and integrin αvβ3. PGB-0-ol also showed an improved ADME profile because of its higher water solubility than PGB-0. PGB-0-ol was synthesized by selective ketone reduction of PGB-0 into primary alcohol by sodium borohydrate producing 30% yield. The cytotoxicity of PGB-0-ol against several breast cancer cells was lower than that of PGB-0. The novel compound PGB-0-ol was synthesized using simple steps. PGB-0-ol has low cytotoxicity against breast cancer cells and could be applied in boron neutron capture therapy as a boron carrier.

Anti‐ Vibrio activity of Pseudoalteromonas xiamenensis STKMTI.2, a new potential vibriosis biocontrol bacterium in marine aquaculture

We isolated Pseudoalteromonas xiamenensis STKMTI.2 from the mangrove sediments of Setokok Island, Indonesia. This study aimed to identify the bacterium, investigate antagonistic properties and minimum inhibitory concentration, and detect the active compounds using gas chromatography–mass spectrometry. The bacterium was identified using a biochemical test and 16S rDNA sequence analysis. Bioactive compounds were extracted from culture supernatant using EtOAc and examined for antibacterial activities against Vibrio spp. on Zobell 2216E agar medium. The antagonistic test of STKMTI.2 against Vibrio harveyi was conducted for 48 h. Active extracts were purified using preparative layer chromatography and tested for MIC. The anti-Vibrio compound was analysed using GCMS. This bacterium was found to be closest to P. xiamenensis Y2T, based on the 16S rDNA sequencing analysis. Anti-Vibrio compound was produced after 96 h of incubation. Strain STKMTI.2 inhibited 50% of the pathogenic Vibrio growth in the co-culture test. The substances in the partially purified sample detected by the GCMS analysis included triethoxy-borane; 1,3-diphenyl-1,3,5,5-tetramethyl-cyclotrisiloxane; 1,6-diazaspiro (4.4) nonane-2,7-dione; and 1,3-bis (4-methoxyphenyl)-5-phenyl-1,3,5-triazinane-2-thione. These substances showed much higher activity against Vibrio spp. compared with commercial antibiotics having MIC ≤0.78 µg ml−1. This study provides a new report on the anti-Vibrio activity of P. xiamenensis STKMTI.2.

Preparation and Cytotoxic Evaluation of PGV-1 Derivative, CCA-1.1, as a New Curcumin Analog with Improved-Physicochemical and Pharmacological Properties.

Purpose: This study aimed to challenge the anticancer potency of pentagamavunone-1 (PGV- 1) and obtain a new compound (Chemoprevention-Curcumin Analog 1.1, CCA-1.1) with improved chemical and pharmacological properties. Methods: CCA-1.1 was prepared by changing the ketone group of PGV-1 into a hydroxyl group with NaBH4 as the reducing agent. The product was purified under preparative layer chromatography and confirmed with HPLC to show about 93% purity. It was tested for its solubility, stability, and cytotoxic activities on several cancer cells. The structure of the product was characterized using 1HNMR, 13C-NMR, FT-IR, and HR-mass spectroscopy. Results: Molecular docking analysis showed that CCA-1.1 performed similar or better interaction to NF-κB pathway-related signaling proteins (HER2, EGFR, IKK, ER-alpha, and ER-beta) and reactive oxygen species (ROS) metabolic enzymes (NQO1, NQO2, GSTP1, AKC1R1, and GLO1) compared with PGV-1, indicating that CCA-1.1 exhibits the same or better anticancer activity than PGV-1. CCA-1.1 also showed better solubility and stability than PGV-1 in aqueous solution at pH 1.0–7.4 under light exposure at room temperature. The cytotoxic activities of CCA-1.1 against several (10) cancer cell lines revealed the same or better potency than PGV-1. Conclusion: In conclusion, CCA-1.1 performs better chemical and anticancer properties than PGV-1 and shows promise as an anticancer agent with high selectivity.

The Investigation of Some Phytochemical Compounds Found in Anchusa strigosa L. Grown Naturally in Iraq

Anchusa strigosa L.: Hardy annual biennial or perennial herb, with hairs especially on the leaves., flowers generally regular. Commonly named (Lisan Althour) in Iraq, from Boraginaceae family. The plant contains phenolic acids, flavonoids, alkaloids, sterols, and terpenoids. The Whole plant part defatted with n-hexane for 24 hours. The defatted plant material extracted using absolute methanol by Soxhlet apparatus for 24 hours, the extract fractionated by solvents of different polarity: petroleum ether- chloroform - ethylacetate- and n-butanol respectively. The n-butanol fraction hydrolyzed with 10% HCl for 5 hours by reflex to break down the glycosidic linkage. Rosmarinic acid, caffeic acid, genistein, and silybin were isolated from ethyl acetate fraction by preparative layer chromatography which identified by high performance liquid chromatography HPLC, Fourier transforms infrared (FTIR) spectra, thin- layer chromatography TLC and melting point. Since the plant contain alkaloids so acid- base extraction performed for crude extract resulting from the maceration of the plant parts in methanol (cold method) to obtain the alkaloid that isolated by preparative layer chromatography and then identified by Fourier transforms infrared (FTIR) spectra and thin-layer chromatography (TLC).
 The aim of this research was to carry out a phytochemical study of this plant since no previous phytochemical investigation work had been done on this species in Iraq.

Detection and isolation of flavonoid and aromatic acid from Cynara scolymus different parts cultivated in iraq

The target of this study was to study the natural phytochemical components of the head (capsule) of Cynara scolymus cultivated in Iraq. The head (capsule) of plant was extracted by maceration in70% ethanol for 72 hours, and fractioned by hexane, chloroform and ethyl acetate. Preliminary qualitative phytochemical screening was performed on the ethyl acetate fraction for capsule was revealed the presence of flavonoid and aromatic acids. These were examined by (high -performance liquid chromatography) (HPLC diodarray), (high- performance thin-layer chromatography)(HPTLC). Flavonoids were isolated by preparative layer chromatography and aromatic acid was isolated by preparative high-performance liquid chromatography HPLC from the ethyl acetate fraction of capsule. Then identified by High Performance Thin Layer Chromatography HPTLC, High performance liquid chromatography HPLC diode array , ultraviolet diode array UV-diode array and Liquid Chromatography /Mass Spectroscopy LC/MS. The chloroform fraction from the capsule was evaluated by Gas Chromatography//Mass Spectrometer(GC/MS). The different chromatographic and spectroscopic techniques revealed the presence of luteolin, apigenin and cinnamic acid in capsule of Cynara scolymus, also 9-octadecanoic acid (oleic acid), Oxalic acid, allyl tetradecyl ester, limonene, in chloroform of Cynara scolymus. The results of the current study proved the presence of luleolin, apigenin, and cinnamic acid in the ethyl acetate fraction of Cynara scolymus capsule.

Anti-Adipogenic compound from Guazuma ulmifolia Leaf

The purpose of this research was to test the anti-adipogenic activity and to identify the active compound of the leaf of Guazuma ulmifolia. The activity of anti-adipogenic of the compound of G. ulmifolia leaf was tested using primary cultures of Wistar strain white mouse preadipocytes. Isolation of the active compound was carried out in the following way. The powder of G. ulmifolia leaf was defated with n-hexane followed by extraction with 70% ethanol. The extract was concentrated and fractionated by liquid-liquid partition using chloroform and ethyl acetate solvents. The ethyl acetate fraction was loaded into a Vacuum Liquid Chromatography (VLC) followed by a reverse phase preparative layer chromatography to obtain a pure isolate. Isolated compound inhibited preadipocytes proliferation and differentiation. Intensive spectroscopic and spectrometric analysis suggested molecular structure of a tiliroside which agreed with previously reported data. The study successfully revealed the traditional claims of the anti-obesity agent of a leaf of G. ulmifolia through inhibition of preadipocytes proliferation and differentiation.

Phytotoxic and antioxidant effects of dichloromethane fraction of Smilax Brasiliensis Sprengel

Dichloromethane (DCM) fraction and sub-fractions obtained from Smilax brasiliensis leaves were examined in order to determine their phytotoxic and antioxidant effects. The dichloromethane fraction was submitted to a preparative layer chromatography leading to seven sub-fractions (DCM1-DCM7). Gas chromatography-mass spectrometry (GC-MS) was performed on the dichloromethane sub-fractions. The DCM sub-fractions presented phytotoxic potential; at a concentration of 125 µg per plate, DCM6 and DCM4 showed the strongest results on Lactuca sativa and Allium cepa, respectively. The DCM fraction and DCM4 sub-fraction were more effective than 2,6-di-tert-butyl-4-methylphenol (BHT) at scavenging the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical. Analysis by GC-MS showed the presence of methyl palmitate (33.05%) in DCM4 and methyl palmitate (17.29%) and methyl oleate (50.96%) in DCM6, suggesting that the activities exhibited by the sub-fractions may be attributed, at least partially, to these major compounds. These results indicate that the DCM sub-fractions of S. brasiliensis could be used as natural herbicides and antioxidants.

Biennial Review of Planar Chromatography: 2015-2017.

The most important advances in planar chromatography published between November 1, 2015, and November 1, 2017, are reviewed in this paper. Included are an introduction to the current status of the field; student experiments and reviews; apparatus and techniques for sample preparation and TLC separations; detection and identification of separated zones; quantitative analysis; preparative layer chromatography; and thin-layer radiochromatography. Selected applications are given in the various sections of the review.

Bioprofiling of Salvia miltiorrhiza via planar chromatography linked to (bio)assays, high resolution mass spectrometry and nuclear magnetic resonance spectroscopy

An affordable bioanalytical workflow supports the collection of data on active ingredients, required for the understanding of health-related food, superfood and traditional medicines. Targeted effect-directed responses of single compounds in a complex sample highlight this powerful bioanalytical hyphenation of planar chromatography with (bio)assays. Among many reports about biological properties of Salvia miltiorrhiza Bunge root (Danshen) and their analytical methods, the highly efficient direct bioautography (DB) workflow has not been considered so far. There was just one TLC-acetylcholinesterase (AChE) method with a poor zone resolution apart from our two HPTLC-DB studies, however, all methods were focused on the nonpolar extracts of Danshen (tanshinones) only. The current study on HPTLC-UV/Vis/FLD-(bio)assay-HRMS, followed by streamlined scale-up to preparative layer chromatography (PLC)-1H-NMR, aimed at an even more streamlined, yet comprehensive bioanalytical workflow. It comprised effect-directed screening of both, its polar (containing phenolics) and nonpolar extracts (containing tanshinones) on the same HPTLC plate, the biochemical and biological profiling with four different (bio)assays and elucidation of structures of known and unidentified active compounds. The five AChE inhibitors, salvianolic acid B (SAB), lithiospermic acid (LSA) and rosmarinic acid (RA) as well as cryptotanshinone (CT) and 15,16-dihydrotanshinone I (DHTI) were confirmed, but also unidentified inhibitors were observed. In the polar extracts, SAB, LSA and RA exhibited free radical scavenging properties in the 2,2-diphenyl-1-picrylhydrazyl assay. CT, DHTI and some unidentified nonpolar compounds were found active against Gram-positive Bacillus subtilis and Gram-negative Aliivibrio fischeri (LOD 12 ng/band for CT, and 5 ng/band for DHTI). For the first time, the most multipotent unidentified active compound zone in the B. subtilis, A. fischeri and AChE fingerprints of the nonpolar Danshen extract was identified as co-eluted band of 1,2-dihydrotanshinone and methylenetanshinquinone in the ratio of 2:1.

Streamlined structure elucidation of an unknown compound in a pigment formulation

A fast and reliable quality control is important for ink manufacturers to ensure a constant production grade of mixtures and chemical formulations, and unknown components attract their attention. Structure elucidating techniques seem time-consuming in combination with column-based methods, but especially the low solubility of pigment formulations is challenging the analysis. In contrast, layer chromatography is more tolerant with regard to pigment particles. One PLC plate for NMR and FTIR analyses and one HPTLC plate for recording of high resolution mass spectra, MS/MS spectra and for gathering information on polarity and spectral properties were needed to characterize a structure, exemplarily shown for an unknown component in pigment Red 57:1 to be 3-hydroxy-2-naphtoic acid. A preparative layer chromatography (PLC) workflow was developed that used an Automated Multiple Development 2 (AMD 2) system. The 0.5-mm PLC plate could still be operated in the AMD 2 system and allowed a smooth switch from the analytical to the preparative gradient separation. Through automated gradient development and the resulting focusing of bands, the sharpness of the PLC bands was improved. For NMR, the necessary high load of the target compound on the PLC plate was achieved via a selective solvent extraction that discriminated the polar sample matrix and thus increased the application volume of the extract that could maximally be applied without overloading. By doing so, the yield for NMR analysis was improved by a factor of 9. The effectivity gain through a simple, but thoroughly chosen extraction solvent is often overlooked, and for educational purpose, it was clearly illustrated and demonstrated by an extended solvent screening. Thus, PLC using an automated gradient development after a selective extraction was proven to be a new powerful combination for structural elucidation by NMR.

Biennial Review of Planar Chromatography: 2013-2015.

The most important advances in planar chromatography published between November 1, 2013 and November 1, 2015 are reviewed in this paper. Included are an introduction to the current status of the field; student experiments, books, and reviews; apparatus and techniques for sample preparation and TLC separations; detection and identification of separated zones; quantitative analysis; preparative layer chromatography; and thin layer radiochromatography. Selected applications are given in the various sections of the review.

Pre-concentration of trace amount of bisphenol A in water samples by palm leaf ash and determination with high-performance liquid chromatography.

Palm leaf ash was characterized and used as low-cost adsorbent for solid-phase extraction and preconcentration of bisphenol A (BPA) in real water samples. Analysis of BPA was carried out using HPLC involving Eurospher 100-5-C18 (25 cm × 4.5 mm, particle size 5 μm) column and water-acetonitrile (40:60, v/v) as mobile phase. The adsorption was achieved quantitatively at a pH of 6 with elution by 3 mL acetonitrile. The limits of detection and enrichment factor were 0.02 μg L(-1) and 333, respectively. Under optimum conditions the relative standard deviation (RSD) was 2% (n = 10). Comparison of qualification criteria of presented preconcentration procedure with other research indicated that palm leaf ash adsorbent was better than many of the adsorbents in terms of cost and reusability. Also, the limit of detection, precision and enrichment factor were comparable and even better than the previously reported methods. Finally, the efficiency of method was computed by determination of trace amounts of BPA in sea, river, mineral and tap waters with recoveries of 93.3-105.5% and RSDs of 0.61-3.12%.. Briefly, the developed solid-phase extraction and Preparative layer chromatography (PLC) methods may be used for bisphenol A monitoring in any environmental water sample. Copyright © 2016 John Wiley & Sons, Ltd.

Biennial Review of Planar Chromatography: 2013-2015.

The most important advances in planar chromatography published between November 1, 2013 and November 1, 2015 are reviewed in this paper. Included are an introduction to the current status of the field; student experiments, books, and reviews; apparatus and techniques for sample preparation and TLC separations; detection and identification of separated zones; quantitative analysis; preparative layer chromatography; and thin layer radiochromatography. Selected applications are given in the various sections of the review.

Effect-Directed Isolation and Identification of Antibacterial Chelidonium majus L. Alkaloids

The components of alkaloid, methanol and ethyl acetate extracts of Chelidonium majus L. root, leaf and stem were tested for antibacterial activity against Bacillus subtilis and Escherichia coli by thin-layer chromatography coupled with direct bioautography (TLC-DB). Effect-directed analyses were performed to obtain the active compounds as follows: separation with preparative layer chromatography (PLC, 0.5-mm-thick silica layer, 20 × 10 cm); a part of the preparative layer (3 cm wide) was used for PLC-DB to detect the antibacterial components against B. subtilis; elution of the appropriate lanes as indicated by PLC-DB from the remaining 17-cm-wide layer; verification of the bioactivity of the eluted compounds by TLC-DB; further purification of the fractions was carried out with TLC, when it was necessary. The eluates of interest were analysed by LC-HRMS and LC-Q/TOF. The antibacterial effect against B. subtilis was mainly attributed to the isoquinoline alkaloid components, which were identified as chelidonine, sanguinarine, chelerythrine, berberine, coptisine, corydine, dihydrosanguinarine, dihydrochelerythrine, 6-acetonyl-dihydrosanguinarine, 6-acetonyl-dihydrochelerythrine and stylopine. Only chelidonine, sanguinarine and chelerythrine possessed inhibiting effect against E. coli.