A fast and validated chromatographic method for simultaneous determination of deferoxamine and Dpenicillamine via chelate formation with metal ions in bulk and dosage forms

Purpose: To develop a chromatographic method for the determination of deferoxamine (DFX) and Dpenicillamine (D-PEN) by improving ultra violet (UV)-absorption via complex formation with Fe2+ and Cu2+ metal ions.Methods: Chromatographic analysis was performed by Waters RP-HPLC system using a Symmetry® C (18) column with a mobile phase comprising 0.1 % formic acid and methanol (95:5 v/v). For complexation process, drug and metal ion solution were mixed in a ratio of 1:5 and the resulting complex directly analyzed. Validation and system suitability parameters (including chromatographic parameters) were determined.Results: DFX-Fe2+ and D-PEN-Cu2+ complexes showed good UV absorption at 260 nm and were easily determined by the newly developed HPLC method. The developed method showed linearity over the concentration range of 8 - 96 μgmL-1 (R2 > 0.999 for DFX and > 0.99 for D-PEN). Precision and accuracy were also within acceptable limits (100.0 ± 2.0 %).Conclusion: The developed method is robust and validated, and satisfies all the system suitability requirements as per ICH guidelines. DFX injection and D-PEN capsule dosage forms can be successfully analysed with the proposed method. The method is simple, fast and cost-effective for the analysis of D-PEN and DFX individually, or simultaneously in bulk drugs as well as in capsule and parenteral formulations, using UV-detector.Keywords: Deferoxamine, D-penicillamine, Chelate formation, Metal ions, HPLC, Dosage form

Thermophoresis for characterizing biomolecular interaction

The study of biomolecular interactions is crucial to get more insight into the biological system. The interactions of protein-protein, protein-nucleic acids, protein-sugars, nucleic acid-nucleic acids and proteinsmall molecules are supporting therapeutics and technological developments. Recently, the development in a large number of analytical techniques for characterizing biomolecular interactions reflect the promising research investments in this field. In this review, microscale thermophoresis technology (MST) is presented as an analytical technique for characterizing biomolecular interactions. Recent years have seen much progress and several applications established. MST is a powerful technique in quantitation of binding events based on the movement of molecules in microscopic temperature gradient. Simplicity, free solutions analysis, low sample volume, short analysis time, and immobilization free are the MST advantages over other competitive techniques. A wide range of studies in biomolecular interactions have been successfully carried out using MST, which tend to the versatility of the technique to use in screening binding events in order to save time, cost and obtained high data qualit

Antidepressant effect of methanol extract of smokeless tobacco and identification of its bioactive components

Purpose: To investigate the antidepressant effect of methanol extract of smokeless tobacco and identify its bioactive compounds. Methods: Adult Wistar rats were randomly assigned to five groups of five rats each: normal control group, standard (reference) control group as well as 100, 200 and 500 mg/kg extract group. The extract, standard drug (imipramine) and normal saline were administered via the intraperitoneal (i.p.) route. The rats were subjected to forced swim test (FST) and tail suspension test (TST) to assess the antidepressant effect of methanol extract of smokeless tobacco. Gas chromatography–mass spectrometry (GC-MS) was used to identify the bioactive compounds of the extract. Results: The oral LD50 of the extract was > 2000 mg/kg. Significant decrease in immobility time was observed after single administration of imipramine (p < 0.05). The extract significantly and dosedependently decreased the immobility time, but increased climbing and swimming times, when compared with normal control group (p < 0.05). The immobility time of stressed rats regardless of sex was significantly and dose-dependently lowered, relative to normal control group (p < 0.05). Four major compounds were identified in the extract: nicotine (45.88 %); 1,5-dimethyl-2-pyrrolidinone (23.00 %), nhexadecanoic acid (11.31 %) and vitamin A aldehyde (9.38 %). Conclusion: These results demonstrate that the methanol extract of smokeless tobacco possesses antidepressant and mood-elevating effects in rats. However, its use should be discouraged since it contains a number of hazardous and carcinogenic components such as N-nitroso compounds and benzo(a)pyrene which are categorized as Class-I carcinogens

Comparing monolithic and fused core HPLC columns for fast chromatographic analysis of fat soluble vitamins

HPLC stationary phases of monolithic and fused core type can be used to achieve fast chromatographic separation as an alternative to UPLC. In this study, monolithic and fused core stationary phases are compared for fast separation of four fat-soluble vitamins. Three new methods on the first and second generation monolithic silica RP-18e columns and a fused core pentafluoro-phenyl propyl column were developed. Application of three fused core columns offered comparable separations of retinyl palmitate, DL-α-tocopheryl acetate, cholecalciferol and menadione in terms of elution speed and separation efficiency. Separation was achieved in approx. 5 min with good resolution (Rs >5) and precision (RSD ≤ 0.6 %). Monolithic columns showed, however, a higher number of theoretical plates, better precision and lower column backpressure than the fused core column. The three developed methods were successfully applied to separate and quantitate fat-soluble vitamins in commercial products

Ultrafast monolithic HPLC method for simultaneous quantification of the anticancer agents, imatinib and sorafenib: Application to tablet dosage forms

Purpose: To develop and validate a simple ultrafast monolithic high performance liquid chromatography (HPLC) method for the simultaneous quantification of two anti-cancer agents, imatinib and sorafenib, in pure form and tablet preparations.Method: Chromatographic separation was accomplished using Chromolith flash RP-18 HPLC-column (25 - 4.6 mm; macropores, 2 μm; mesopores, 13 – 15 nm). The optimum mobile phase composition of ammonium acetate buffer (10 mM, pH 8.5) and methanol at ratio of 35:65 v/v was used. Effluent flow rate was adjusted to 1.0 mL/min and the analysis was performed at 250 nm wavelength. The developed method was evaluated for specificity, linearity, precision and accuracy.Results: The method offered a linear relationship over the concentration range of 1 - 16 μg/ml (correction coefficient, R2 = 0.9999) for both analytes. Limit of detection (LOD) was 0.1891 and 0.1888 μg/ml while limit of quantification (LOQ) was 0.6303 and 0.6294 μg/ml for imatinib and sorafenib, respectively. Mean recovery was within 100 ± 2 %. The utility of the new method was demonstrated by its successful use for the analysis of commercially available tablet formulations of both drugs.Conclusion: The developed method is fast and economical, and is being recommended for routine analysis of imatinib and sorafenib in bulk drug and tablet dosage forms in quality control laboratories.Keywords: RP-HPLC, Chromolith, Imatinib, Sorafenib, Validation, Quality contro

Methanol extract of smokeless tobacco alters inflammation and nociception process in animal models

Purpose: To investigate the inflammatory and nociceptive alterations due to the use of Nicotiana tabacum or smokeless tobacco (MEST) owing to the fact that it is used by some people to relieve dental pain.Methods: Hepatic biochemical indicators and thiobarbituric acid reactive substances assay were used to assess MEST toxicity and pharmacological doses selection. The effects on inflammation of different pharmacological doses (100, 200 and 500 mg/kg i.p.) of MEST were evaluated using xylene-induced ear edema and cotton pellet granuloma tests. Indomethacin (10 mg/kg, i.p.) was used as positive standard drug, whereas the vehicle 0.5 % CMC treated group was considered as negative control. Acetic acid-induced abdominal contraction test and formalin-induced hind paw licking model were utilized to assess the role of MEST in nociception. Indomethacin (10 mg/kg i.p.) and diclofenac sodium (10 mg/kg i.p.) were used as positives standard drugs. The vehicle used was 0.5% CMC which served as the negative control.Results: MEST (50 %, 200 mg/kg) and indomethacin (47.5 %) both elicited a significant (p < 0.001) anti-edematogenic effect on xylene-induced ear edema. MEST also showed a significant (p < 0.001) inhibitory effect on granuloma formation at all administered doses as compared to the untreated groups which was comparable to standard drug indomethacin. The number of acetic acid induced writhings was observed to be significantly increased (p < 0.001) by MEST at all doses, unlike diclofenac that led to significant reduction (p < 0.001) in the number of writhings, when compared to the untreated group. MEST also showed a significant (p < 0.05) dose-dependent reduction of the hind paw licking caused by formalin when compared to the vehicle control.Conclusion: These results signify that administration of MEST induces inflammatory and nociceptive alterations. However, the extract is not recommended for dental pain due to its other toxic effects that have previously been reported.Keywords: Nicotiana tabacum, Smokeless tobacco, Inflammation, Nociceptio

Investigation of the presence of pharmaceuticals and personal care products (PPCPs) in groundwater of Jazan area, Saudi Arabia

Purpose: To investigate the possible occurrence of some selected  pharmaceutical compounds in the groundwater of Jazan area, Saudi Arabia.Methods: Water samples from 46 wells were collected from different sites covering Jazan area of Saudi Arabia between February and March 2017. These samples were first analyzed to investigate the presence of eleven drugs mostly used in the study area. Thereafter, samples were subjected to liquid chromatography-mass spectrometry (LC-MS/MS) by direct injection and external standard calibration.Results: Despite the low detection limit (0.001 - 0.02 μg/L) applied to the investigated compounds with a variety chemical groups (acetylsalicylic acid, paracetamol, ibuprofen, metronidazole, caffeine, olmesartan, omeprazole, nifedipine, diclofenac sodium, glibenclamide and loratidine), none of these compounds was detected in any of the analyzed samples.Conclusion: The main source of environmental contamination with  pharmaceuticals and personal care products (PPCPs) is wastewater. The results obtained reveal the absence of groundwater contamination by these compounds in Jazan area. However, further extended investigations and monitoring are recommended.Keywords: Pharmaceuticals, Groundwater, Wastewater, Pollution, Personal care products, Liquid chromatography-mass spectrometer (LC-MS/MS

Phytochemical, antimicrobial and cytotoxicity screening of ethanol extract of Acacia ehrenbergiana Hayne grown in Jazan Region of Saudi Arabia

Purpose: To explore the phytoconstituents of Acacia ehrenbergiana Hayne as well as its biological effects. Methods: Determination of phytoconstituents of ethanol extract of the plant was performed by gas chromatography-mass spectrometry (GC-MS) technique. Antibacterial screening was conducted against the isolates of Gram-positive and Gram-negative microbes while the anti-carcinogenic properties of the ethanol extract on cancerous cells were investigated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cytotoxicity assay against breast MCF7, ovary cancer A2780 and colon cancer HT29 cells, respectively, in addition to normal MRC5 fibroblast cells. Results: GC-MS analysis identified 15 different phytochemicals in the ethanol extract. The extract exerted significant antimicrobial activity with the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) in the range 1.56 - 6.25 and 3.12 – 12.5 mg/L, respectively, against all test bacterial strains. Cytotoxic activity, obtained by MTT assay, was 28.81 ± 0.99, 12.50 ± 2.50, 23.90 ± 0.74 and 50.58 ± 3.24 μg/mL, against the three cancer cell lines and normal fibroblast, respectively. MTT cytotoxicity results was further confirmed by clonogenic survival assay on MCF7 cells. Conclusion: This study highlights the potential interesting ethnopharmacological applications of Acacia ehrenbergiana Hayne to treat drug-resistant pathogens as standardized extract. Keywords: Acacia ehrenbergiana, Phytochemistry, Antimicrobial, Cytotoxicit

High performance thin-layer chromatography and in vitro cytotoxic studies on ethanol extract of Matricaria chamomilla L (Asteraceae) flowers

Purpose: To develop a high performance thin-layer chromatography (HPTLC procedure for quantitation of apigenin in ethanol extract of Matricaria chamomilla (Babunaj) flowers, and to evaluate the extract for in vitro cytotoxic effect on MCF-7 cell lines. Methods: Quantification of apigenin was carried out using a CAMAG TLC system. A combination of toluene, ethyl acetate and formic acid (4.5:3.5:0.2 v/v/v) was used as mobile phase, with densitometry detection at 336 nm. The HPTLC procedure was subjected to validation as per ICH guidelines. The cytotoxicity of the extract was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Results: A sharp apigenin band at Rf of 0.51 was obtained, and the content of apigenin in the extract was 0.062 % w/w. The detection limit (LOD) and quantification limit (LOQ) were 0.19 and 0.57 ng/band, respectively. MTT assay results indicate that M. chamomilla was cytotoxic to Michigan Cancer Foundation-7 (MCF-7) cells, with half-maximal concentration (IC50) of 74 µg/mL. Conclusion: The developed HPTLC method is linear, precise, accurate and specific for the determination of apigenin. M. chamomilla exerts cytotoxic effect on MCF-7 cell line via induction of apoptosis

Polymorphism of HLA and Susceptibility of Breast Cancer

Breast cancer (BC) is the second most common malignancy in the world. Numerous studies have demonstrated the association between human leukocyte antigen (HLA) and cancer. The occurrence and development of BC are closely linked to genetic factors. Human leukocyte antigens G and E (HLA-G and HLA-E) are non-classical major histocompatibility complex (MHC) class I molecules. These molecules play an important role in immune surveillance by inhibiting the cytotoxic and natural killer T cells responsible for immune escape. The expression of HLA-G and HLA-E has been associated with several diseases, including tumors. The HLA system plays a key role in the escape of tumor cells from immune surveillance. This review aims to determine the correlation between BC susceptibility and HLA markers specific HLA alleles such as HLA-B07, HLA-DRB111, HLA-DRB113, and HLA-DRB115 are associated with an increased risk of developing BC. Furthermore, HLA-G mutations have been attributed to an elevated likelihood of metastasis in BC patients. Understanding the complex associations between the HLA system and BC development is critical for developing novel cancer prevention, detection, and treatment strategies. This review emphasizes the importance of analyzing HLA polymorphisms in the management of BC patients, as well as the urgent need for further research in this area