Silica Gel TLC Plates Research Articles

Thin-layer chromatography of polar and ionic compounds using active gas flow over the silica gel adsorbent layer

Separation of polar and ionic substances in carbon dioxide and ammonia atmospheres on unmodified silica gel TLC plates with reduced moisture content has been studied by single and multiple development chromatography with countercurrent drying. Regulation of moisture content and of saturation of the layer with mobile phase vapor, creation of active atmospheres, and countercurrent drying of the mobile phase was achieved by flow of gas over the adsorbent layer. The possibility of simple gradient separation by passage of polar vapor is also shown.

Cloning and Characterization of an Armillaria gallica cDNA Encoding Protoilludene Synthase, Which Catalyzes the First Committed Step in the Synthesis of Antimicrobial Melleolides

Melleolides and related fungal sesquiterpenoid aryl esters are antimicrobial and cytotoxic natural products derived from cultures of the Homobasidiomycetes genus Armillaria. The initial step in the biosynthesis of all melleolides involves cyclization of the universal sesquiterpene precursor farnesyl diphosphate to produce protoilludene, a reaction catalyzed by protoilludene synthase. We achieved the partial purification of protoilludene synthase from a mycelial culture of Armillaria gallica and found that 6-protoilludene was its exclusive reaction product. Therefore, a further isomerization reaction is necessary to convert the 6–7 double bond into the 7–8 double bond found in melleolides. We expressed an A. gallica protoilludene synthase cDNA in Escherichia coli, and this also led to the exclusive production of 6-protoilludene. Sequence comparison of the isolated sesquiterpene synthase revealed a distant relationship to other fungal terpene synthases. The isolation of the genomic sequence identified the 6-protoilludene synthase to be present as a single copy gene in the genome of A. gallica, possessing an open reading frame interrupted with eight introns.

Catalytic Reactions of the Homogentisate Prenyl Transferase Involved in Plastoquinone-9 Biosynthesis

Homogentisate solanesyl transferase (HST) catalyzes the prenylation and decarboxylation of homogentisate to form 2-methyl-6-solanesyl-1,4-benzoquinol, the first intermediate in plastoquinone-9 biosynthesis. In vitro, HST from Spinacia oleracea L., Arabidopsis thaliana, and Chlamydomonas reinhardtii were all found to use not only solanesyl diphosphate but also short chain prenyl diphosphates of 10-20 carbon atoms as prenyl donors. Surprisingly, with these donors, prenyl transfer was largely decoupled from decarboxylation, and thus the major products were 6-prenyl-1,4-benzoquinol-2-methylcarboxylates rather than the expected 2-methyl-6-prenyl-1,4-benzoquinols. The 6-prenyl-1,4-benzoquinol-2-methylcarboxylates were not substrates for HST-catalyzed decarboxylation, and the enzyme kinetics associated with forming these products appeared quite distinct from those for 2-methyl-6-prenyl-1,4-benzoquinol formation in respect of catalytic rate, substrate K(m) value, and the pattern of inhibition by haloxydine, a molecule that appeared to act as a dead end mimic of homogentisate. These observations were reconciled into a simple model for the HST mechanism. Here, prenyl diphosphate binds to HST to form at least two alternative complexes that go on to react differently with homogentisate and prenylate it either with or without it first being decarboxylated. It is supposed that solanesyl diphosphate binds tightly and preferentially in the mode that compels prenylation with decarboxylation.

Amplification of Adenine Phosphoribosyltransferase Suppresses the Conditionally Lethal Growth and Virulence Phenotype of Leishmania donovani Mutants Lacking Both Hypoxanthine-guanine and Xanthine Phosphoribosyltransferases

Leishmania donovani cannot synthesize purines de novo and obligatorily scavenge purines from the host. Previously, we described a conditional lethal Deltahgprt/Deltaxprt mutant of L. donovani (Boitz, J. M., and Ullman, B. (2006) J. Biol. Chem. 281, 16084-16089) that establishes that L. donovani salvages purines primarily through hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and xanthine phosphoribosyltransferase (XPRT). Unlike wild type L. donovani, the Deltahgprt/Deltaxprt knock-out cannot grow on 6-oxypurines and displays an absolute requirement for adenine or adenosine and 2'-deoxycoformycin, an inhibitor of parasite adenine aminohydrolase activity. Here, we demonstrate that the ability of Deltahgprt/Deltaxprt parasites to infect mice was profoundly compromised. Surprisingly, mutant parasites that survived the initial passage through mice partially regained their virulence properties, exhibiting a >10-fold increase in parasite burden in a subsequent mouse infection. To dissect the mechanism by which Deltahgprt/Deltaxprt parasites persisted in vivo, suppressor strains that had regained their capacity to grow under restrictive conditions were cloned from cultured Deltahgprt/Deltaxprt parasites. The ability of these suppressor clones to grow in and metabolize 6-oxypurines could be ascribed to a marked amplification and overexpression of the adenine phosphoribosyltransferase (APRT) gene. Moreover, transfection of Deltahgprt/Deltaxprt cells with an APRT episome recapitulated the suppressor phenotype in vitro and enabled growth on 6-oxypurines. Biochemical studies further showed that hypoxanthine, unexpectedly, was an inefficient substrate for APRT, evidence that could account for the ability of the suppressors to metabolize hypoxanthine. Subsequent analysis implied that APRT amplification was also a potential contributory mechanism by which Deltahgprt/Deltaxprt parasites displayed persistence and increased virulence in mice.

AKTIVITAS INSEKTISIDA EKSTRAK DAUN DAN BIJI TEPHROSIA VOGELII J. D. HOOKER (LEGUMINOSAE) DAN EKSTRAK BUAH PIPER CUBEBA L. (PIPERACEAE) TERHADAP LARVA CROCIDOLOMIA PAVONANA (F.) (LEPIDOPTERA: CRAMBIDAE)

Insecticidal activity of leaf and seed extracts of Tephrosia vogelii J. D. Hooker (Leguminosae) and fruit extract of Piper cubeba L. (Piperaceae) on the cabbage head caterpillar, Crocidolomia pavonana (L.) (Lepidoptera: Crambidae). Ethyl acetate leaf and seed extracts of Tephrosia vogelii and a solid fraction of ethyl acetate fruit extract of Piper cubeba were evaluated for their insecticidal activity on second-instar larvae Crocidolomia pavonana by a leaf-residue feeding method in the laboratory. Leaf extracts of purple and white-flowered T. vogelii showed the same pattern of component separation on silica gel TLC plate (Rf between 0.21 and 0.94), and likewise the separation of components of seed extracts of purple and white-flowered T. vogelii (Rf between 0.31 and 0.96). All four kinds of T. vogelii extracts showed intense UV-absorbing nonpolar spots (Rf > 0.8). Based on LC50 ratio at day 4, leaf extract of purple-flowered T. vogelii (LC50 0.075%) was 4.30, 2.70, 2.21, and 1.64 times more toxic than fruit extract of P. cubeba, seed extract of white-flowered T. vogelii, seed extract of purple-flowered T. vogelii, and leaf extract of white-flowered T. vogelii, respectively. All T. vogelii extracts were more toxic to C. pavonana larvae than P. cubeba fruit extract. At LC95 level, a mixture of leaf extract of purple-flowered T. vogelii and fruit extract of P. cubeba (5:9, w/w) was more toxic to C. pavonana larvae than each extract tested separately. This extract mixture had synergistic joint action against C. pavonana larvae both at LC50 and LC95 level. Thus, leaf extract of purple-flowered T. vogelii and its mixture with P. cubeba fruit extract are promising to be used for controlling C. pavonana.

Rapid Exchange of Bound ADP on the Staphylococcus aureus Replication Initiation Protein DnaA

In Escherichia coli, regulatory inactivation of the replication initiator DnaA occurs after initiation as a result of hydrolysis of bound ATP to ADP, but it has been unknown how DnaA is controlled to coordinate cell growth and chromosomal replication in gram-positive bacteria such as Staphylococcus aureus. This study examined the roles of ATP binding and its hydrolysis in the regulation of the S. aureus DnaA activity. In vitro, S. aureus DnaA melted S. aureus oriC in the presence of ATP but not ADP by a mechanism independent of ATP hydrolysis. Unlike E. coli DnaA, binding of ADP to S. aureus DnaA was unstable. As a result, at physiological concentrations of ATP, ADP bound to S. aureus DnaA was rapidly exchanged for ATP, thereby regenerating the ability of DnaA to form the open complex in vitro. Therefore, we examined whether formation of ADP-DnaA participates in suppression of replication initiation in vivo. Induction of the R318H mutant of the AAA+ sensor 2 protein, which has decreased intrinsic ATPase activity, caused over-initiation of chromosome replication in S. aureus, suggesting that formation of ADP-DnaA suppresses the initiation step in S. aureus. Together with the biochemical features of S. aureus DnaA, the weak ability to convert ATP-DnaA into ADP-DnaA and the instability of ADP-DnaA, these results suggest that there may be unidentified system(s) for reducing the cellular ratio of ATP-DnaA to ADP-DnaA in S. aureus and thereby delaying the re-initiation of DNA replication.

Chromatographic-densitometric Method for Determination of Clopamide and 4-chlorobenzoic, and 4-chloro-3-sulfamoylbenzoic Acids in Tablets

New chromatographic-densitometric method for determination of clopamide and impurities (4-chlorobenzoic and 4-chloro-3-sulfamoylbenzoic acids) in tablets was developed. Silica gel TLC plates as a stationary phase and nbutanol- 2-propanol -water- methylene chloride (10:7:2:5:3 v/v/v/v/v) as a mobile phase were used for separation. Densitometric measurements were done at λ = 235 nm. Favourable retention parameters (Rf, Rs, k, α) were obtained under developed conditions, which guarantee good separation of studied components. Whereas, results that were obtained from validation method confirm specificity, high sensitivity, linearity in a range of studied concentrations, repeatability and good accuracy of this method.

Simultaneous Identification and Quantitative Determination of Selected Aminoglycoside Antibiotics by Thin-Layer Chromatography and Densitometry

A TLC-densitometric method has been developed for simultaneous identification and quantitative determination of amikacin, gentamicin, kanamycin, neomycin, netilmicin, and tobramycin. This separation of antibiotics was achieved on silica gel TLC plates without a fluorescent indicator and with methanol-25% ammonia-chloroform (3 + 2 + 1, v/v/v) as the mobile phase. The densitometric measurements were made at 500 nm after detection with a 0.2% ninhydrin solution in ethanol. Under these conditions, good separation of the chosen aminoglycosides was obtained. The method is distinguished by high sensitivity, with the LOD from 0.25 microg for amikacin to 1.00 microg for gentamicin and the LOQ from 0.5 microg for amikacin to 1.65 microg for gentamicin, and a wide linearity range 0.75-6.25 microg/spot for amikacin and netilmicin and 1.5-12.50 microg/spot for other antibiotics. The precision of the determination was very good; RSD varied in the range 0.3-0.6%.

Evidence for a Second, High Affinity Gβγ Binding Site on Gαi1(GDP) Subunits

It is well known that Galpha(i1)(GDP) binds strongly to Gbetagamma subunits to form the Galpha(i1)(GDP)-Gbetagamma heterotrimer, and that activation to Galpha(i1)(GTP) results in conformational changes that reduces its affinity for Gbetagamma subunits. Previous studies of G protein subunit interactions have used stoichiometric amounts of the proteins. Here, we have found that Galpha(i1)(GDP) can bind a second Gbetagamma subunit with an affinity only 10-fold weaker than the primary site and close to the affinity between activated Galpha(i1) and Gbetagamma subunits. Also, we find that phospholipase Cbeta2, an effector of Gbetagamma, does not compete with the second binding site implying that effectors can be bound to the Galpha(i1)(GDP)-(Gbetagamma)(2) complex. Biophysical measurements and molecular docking studies suggest that this second site is distant from the primary one. A synthetic peptide having a sequence identical to the putative second binding site on Galpha(i1) competes with binding of the second Gbetagamma subunit. Injection of this peptide into cultured cells expressing eYFP-Galpha(i1)(GDP) and eCFP-Gbetagamma reduces the overall association of the subunits suggesting this site is operative in cells. We propose that this second binding site serves to promote and stabilize G protein subunit interactions in the presence of competing cellular proteins.

Separation and determination of rofecoxib and its degradation products by TLC chromatography

A thin layer chromatography with densitometry method for the determination of rofecoxib and degradation products is described. The chromatographic separation was performed on silica gel TLC plates as a stationary phase and chloroform-acetone-toluene-glacial acetic acid (12: 5: 2: 0.1, v/v/v/v) as a mobile phase. Densitimetric detection was carried out at 256 nm. The method is of high sensitivity and low LOD and LOQ: from 0.35 μg/spot to 1.05 μg/spot. The recovery was satisfactory at 98.62%. In addition, the stability of rofecoxib in solutions was investigated, including an effect of solution pH, temperature and incubation time. The method is rapid, easy and selective, particularly for the analysis of rofecoxib formulations.

Enantioseparation of cetirizine by chromatographic methods and discrimination by 1H‐NMR

Cetirizine is an antihistaminic drug used to prevent and treat allergic conditions. It is currently marketed as a racemate. The H1-antagonist activity of cetirizine is primarily due to (R)-levocetirizine. This has led to the introduction of (R)-levocetirizine into clinical practice, and the chiral switching is expected to be more selective and safer. The present work represents three methods for the analysis and chiral discrimination of cetirizine. The first method was based on the enantioseparation of cetirizine on silica gel TLC plates using different chiral selectors as mobile phase additives. The mobile phase enabling successful resolution was acetonitrile-water 17: 3, (v/v) containing 1 mM of chiral selector, namely hydroxypropyl-beta-cyclodextrin, chondroitin sulphate or vancomycin hydrochloride. The second method was a validated high performance liquid chromatography (HPLC), based on stereoselective separation of cetirizine and quantitative determination of its eutomer (R)-levocetirizine on a monolithic C18 column using hydroxypropyl-beta-cyclodextrin as a chiral mobile phase additive. The resolved peaks of (R)-levocetirizine and (S)-dextrocetirizine were confirmed by further mass spectrometry. The third method used a (1)H-NMR technique to characterize cetirizine and (R)-levocetirizine. These methods are selective and accurate, and can be easily applied for chiral discrimination and determination of cetirizine in drug substance and drug product in quality control laboratory. Moreover, chiral purity testing of (R)-levocetirizine can also be monitored by the chromatographic methods.

Simple densitometric TLC analysis of plaunotol for screening of high-plaunotol-containing Plants ofCroton stellatopilosusOhba

Plaunotol, an anti-peptic diterpenoid present in the leaves of Croton stellatopilosus Ohba, was separated from other components of crude methanolic extracts and quantified by densitometric TLC. Validation of precision, repeatability, and accuracy showed the technique was reliable, with 98.7% recovery of externally added plaunotol. Crude extracts of different leaf samples could be applied directly to silica gel TLC plates without prior sample preparation or concentration of the extracts. Plates were chromatographed with chloroform-n-propanol 96:4 (v/v) as mobile phase then scanned at 220 nm. The results showed that the plaunotol content of the leaves of C. stellatopilosus was highly variable (range 0.14 to 0.79% w/w of dry weight). Although most (35%) leaf samples seemed to contain 0.3–0.4% plaunotol, 8% contained more than 0.6%. This densitometric TLC method could, therefore, be a useful tool for screening of high-plaunotol-producing C. stellatopilosus plants as effective sources of the compound.

Purification and Characterization of the Lipid A 1-Phosphatase LpxE of Rhizobium leguminosarum

LpxE, a membrane-bound phosphatase found in Rhizobium leguminosarum and some other Gram-negative bacteria, selectively dephosphorylates the 1-position of lipid A on the outer surface of the inner membrane. LpxE belongs to the family of lipid phosphate phosphatases that contain a tripartite active site motif and six predicted transmembrane helices. Here we report the purification and characterization of R. leguminosarum LpxE. A modified lpxE gene, encoding a protein with an N-terminal His6 tag, was expressed in Escherichia coli. The protein was solubilized with Triton X-100 and purified to near-homogeneity. Gel electrophoresis reveals a molecular weight consistent with the predicted 31 kDa. LpxE activity is dependent upon Triton X-100, optimal near pH 6.5, and Mg2+-independent. The H197A and R133A substitutions inactivate LpxE, as does treatment with diethyl pyrocarbonate. In a mixed micelle assay system, the apparent Km for the precursor lipid IV(A) is 11 microm. Substrates containing the 3-deoxy-d-manno-oct-2-ulosonic acid disaccharide are dephosphorylated at similar rates to lipid IV(A), whereas glycerophospholipids like phosphatidic acid or phosphatidylglycerol phosphate are very poor substrates. However, an LpxE homologue present in Agrobacterium tumefaciens is selective for phosphatidylglycerol phosphate, demonstrating the importance of determining substrate specificity before assigning the functions of LpxE-related proteins. The availability of purified LpxE will facilitate the preparation of novel 1-dephosphorylated lipid A molecules that are not readily accessible by chemical methods.

Simultaneous Determination of Pyrathiazine Theoclate and Pyridoxine HCl by TLC-Densitometry in Commercial Tablets: Validation of the Method

A simple and rapid densitometric method has been developed for determination of pyrathiazine theoclate and pyridoxine HCl in commercial tablets. After extracting the samples four times with ethanol, the solutions were spotted on pre-coated silica gel TLC plates, which were eluted with a mixture of dichloromethane–methanol–acetone–acetic acid (19.5:6.0:3.0:1.5, v/v). Quantitative evaluation was performed by measuring the absorbance reflectance of the analyte spots at λ = 254 nm. The TLC-densitometric method is cheap, selective, precise, and accurate and can be used for routine analysis of pyrathiazine theoclate and pyridoxine HCl in tablet preparations at industry quality control laboratories.

Determination of Piroxicam and Degradation Products in Drugs by TLC

A thin-layer chromatography method with densitometric detection for the determination of piroxicam and its degradation products is described. Separation was on silica gel TLC plates with ethyl acetate/toluene/butylamine (2+2+1, v/v/v) as the mobile phase. Densitometric detection was carried out at 360 nm. The method has high sensitivity, and satisfactory recovery. Based on 1H NMR and LC–MS–MS, it was found that piroxicam decomposes to produce pyridine-2-amine and 2-methyl-2,3-dihydro-4H-1λ6,2-benzotiazin-1,1,4-trione.

A Trypanosoma cruzi Phosphatidylinositol 3-Kinase (TcVps34) Is Involved in Osmoregulation and Receptor-mediated Endocytosis

Trypanosoma cruzi, the etiological agent of Chagas disease, has the ability to respond to a variety of environmental changes during its life cycle both in the insect vector and in the vertebrate host. Because regulation of transcription initiation seems to be nonfunctional in this parasite, it is important to investigate other regulatory mechanisms of adaptation. Regulatory mechanisms at the level of signal transduction pathways involving phosphoinositides are good candidates for this purpose. Here we report the identification of the first phosphatidylinositol 3-kinase (PI3K) in T. cruzi, with similarity with its yeast counterpart, Vps34p. TcVps34 specifically phosphorylates phosphatidylinositol to produce phosphatidylinositol 3-phosphate, thus confirming that it belongs to class III PI3K family. Overexpression of TcVps34 resulted in morphological and functional alterations related to vesicular trafficking. Although inhibition of TcVps34 with specific PI3K inhibitors, such as wortmannin and LY294,000, resulted in reduced regulatory volume decrease after hyposmotic stress, cells overexpressing this enzyme were resistant to these inhibitors. Furthermore, these cells were able to recover their original volume faster than wild type cells when they were submitted to severe hyposmotic stress. In addition, in TcVps34-overexpressing cells, the activities of vacuolar-H+-ATPase and vacuolar H+-pyrophosphatase were altered, suggesting defects in the acidification of intracellular compartments. Furthermore, receptor-mediated endocytosis was partially blocked although fluid phase endocytosis was not affected, confirming a function for TcVps34 in membrane trafficking. Taken together, these results strongly support that TcVps34 plays a prominent role in vital processes for T. cruzi survival such as osmoregulation, acidification, and vesicular trafficking.

TLC study of the separation of the enantiomers of lactic acid

Summary Enantiomer separation by TLC is still much less frequent than with other, mostly instrumental, chromatographic techniques. From a literature survey it is apparent that separation of the enantiomers of d,l-lactic acid is primarily of interest to the diary industry and that this particular separation is less frequently performed by chromatographic than by membrane techniques. As far as we are aware, before our studies only one report of TLC separation of the enantiomers of d,l-lactic acid was available in the literature; this is dated 1991 and describes the use of non-instrumental TLC only. In this study, we started by reproducing the TLC procedure originating from 1991, for this purpose using TLC with automatic sample application and densitometric detection. We managed to repeat the earlier procedure and to achieve full, i.e. baseline, separation of the enantiomers, with a remarkable distance between the two antimers. However, we revealed a significant drawback of this separation procedure — d-(−)-lactic acid was transported almost with the mobile-phase front and its densitometric quantification was barely possible because of the relatively high UV absorption of the mobile-phase front line. The reference method for separation of the enantiomers of d,l-lactic acid consisted in preliminary impregnation of commercial silica gel TLC plates with copper(II) acetate. In-situ formation of bidentate complexes of the d,l-lactic acid antimers with the Cu2+ cation resulted in different mobilities of these complex cations in the planar chromatographic system. The objectives of this study were twofold — to investigate separation of the enantiomers of d,l-lactic acid with other transition metal cations (i.e., Co2+, Ni2+, and Mn2+) used to impregnate the silica gel (to achieve resolution that might enable quantification of the two lactic acid antimers and not only the l-(+) enantiomer) and to gain deeper insight into the mechanism of separation with these metal cations. For purposes of comparison, we chromatographed d,l-lactic acid on non-impregnated silica gel layers. As a result, we managed to establish efficient separation conditions with the Ni2+ and Co2+ cations that outperformed the earlier established procedure involving the Cu2+ cation, and — partially at least — to elucidate the mechanism of separation of the enantiomers of d,l-lactic acid by these TLC systems. The Mn2+ cation proved unsuitable for the purpose. Finally, we managed to separate the enantiomers of d, l-lactic acid on non-impregnated silica gel layer also, which seems yet more proof of the microcrystalline chirality of silica gel used as stationary phase and of its substantial contribution to the enantiomer separation investigated.

Role of the Diacylglycerol Kinase α-Conserved Domains in Membrane Targeting in Intact T Cells

Diacylglycerol kinase (DGK) phosphorylates diacylglycerol to phosphatidic acid, modifying the cellular levels of these two lipid mediators. Ten DGK isoforms, grouped into five subtypes, are found in higher organisms. All contain a conserved C-terminal domain and at least two cysteine-rich motifs of unknown function. DGKalpha is a type I enzyme that acts as a negative modulator of diacylglycerol-based signals during T cell activation. Here we studied the functional role of the DGKalpha domains using mutational analysis to investigate membrane binding in intact cells. We show that the two atypical C1 domains are essential for plasma membrane targeting of the protein in intact cells but unnecessary for catalytic activity. We also identify the C-terminal sequence of the protein as essential for membrane binding in a phosphatidic acid-dependent manner. Finally we demonstrate that, in the absence of the calcium binding domain, receptor-dependent translocation of the truncated protein is regulated by phosphorylation of Tyr(335). This functional study provides new insight into the role of the so-called conserved domains of this lipid kinase family and demonstrates the existence of additional domains that confer specific plasma membrane localization to this particular isoform.

Optimization of growth conditions for test fungus cultures used in direct bioautographic TLC detection. 3. Test fungus:Candida albicans

Summary Optimum conditions have been established for culture of the fungus Candida albicans (ATCC 90028) for microbial detection of zones in direct bioautographic TLC. Bioluminescent ATP assay is a highly sensitive method for optimizing the viability of Candida albicans test fungus in bioautographic TLC detection. A suspension of microbes (OD 600nm = 0.5–0.7, in Mueller–Hinton broth with 5% glucose) in the log phase of growth can be used for dipping TLC plates. On the basis of our results with Candida albicans, we can differentiate between microbiostatic (bacteriostatic or fungistatic) and microbiocidal (bactericidal or fungicidal) effects on TLC plates. Our micrographs clearly show the borders of inhibition zones in bioautograms. This technique leads to new possibilities in studies of the interactions between microbes and antimicrobial compounds on bioautographic silica gel TLC plates by using scanning electron microscopy. In this paper we describe an optimization procedure for bioautographic TLC detection.

Simultaneous Densitometric Determination of 6‐Gingerol and 6‐Shogaol in some Commercial Gingers (Zingiber Officinale Roscoe)

A simple and rapid densitometric method has been developed for determination of 6‐gingerol and 6‐shogaol in some commercial gingers. After extracting the samples four times with methanol, the solutions were spotted on pre‐coated silica gel tlc plates, which were eluted with a mixture of n‐hexane‐ethyl ether (4:6, v/v). Quantitative evaluation was performed by measuring the absorbance reflectance of the analyte spots at λ=577 nm after sprayed with anisaldehyde‐h2so4 reagent. The tlc‐densitometric method is cheap, selective, precise and accurate and can be used for routine analysis of gingers in herbal drugs industry quality control laboratories.