Spectrophotometric Methods for Determination of Sunitinib in Pharmaceutical Dosage Forms Based on Ion-pair Complex Formation.

Three rapid spectrophotometric methods were developed for the determination of sunitinib based on the formation of ion-pair complex in acidic medium with bromocresol purple, bromothymol blue, and bromophenol blue. The formed ion-pair complexes, extractable with chloroform, were measured at 422 nm for bromocresol purple, 425 nm for bromothymol blue and 427 nm for bromophenol blue. All these methods were optimized for the pH of buffer and the volume of the reagent. The methods were linear over the range of 1-200 µg/mL for bromocresol purple, 1-150 µg/mL for bromothymol blue, and 2-200 µg/mL for bromophenol blue with a very low limit of quantification and acceptable accuracy and precision. Using the proposed methods for determination of sunitinib in pharmaceutical dosage forms showed reliable results comparable to previously published method.

Bioassay-guided fractionation and identification of wound healing active compound from Pistacia vera L. hull extract.

ETHNOPHARMACOLOGICAL RELEVANCE: Pistachio hull has traditionally been used to treat peptic ulcer, hemorrhoids, oral and cutaneous wounds. AIM OF THE STUDY: On the basis of its traditional uses and previous pharmacological reports, a bioassay guided fractionation procedures on pistachio (Pistacia vera L.) hulls was performed to define the fractions and bioactive compound that are responsible for wound healing activity of hulls. MATERIAL AND METHODS: A bioassay-guided fractionation of the total extract (MeOH 80%) of Pistacia vera L. hulls was carried out to evaluate wound healing activity by scratch assay on NIH/3T3 murine fibroblast cells. A combination of solvent-solvent partitioning, column chromatography, preparative thin layer chromatography and crystallization were used to obtain fractions/sub-fractions and pure compound. The wound healing potential of isolated compound was examined by fibroblasts migration and proliferation using scratch assay and CFSC dilution assay, respectively. In addition, we evaluated the gene expression of some inflammatory markers which are involved in healing process using Real Time PCR. Chemical structure of active compound was elucidated by spectrometric methods. RESULTS: Due to the higher wound healing activity of CHCl3 fraction from P. vera hulls, it was fractionated by successive chromatographic techniques to yield the active compound. 3-Epimasticadienolic acid was isolated and crystallized as a white powder. This active compound (200 µg/ml) significantly increased the fibroblast proliferation and migration, resulting in reduction of the scratch area about 45%. It showed a strong inhibitory effect on gene expression of IL-6 and TNF-α, and a stimulation effect on NF-κB gene expression at the same dose. CONCLUSION: The present study supported the traditional uses of P. vera hulls for wound-healing and 3-epimasticadienolic acid showed significantly potent on wound repair.

Evaluation of anti-tyrosinase activity of Allium ursinum extracts and their metal complexes.

BACKGROUND: A. ursinum is found to contain high levels of some beneficial phenolic and poly phenolic compounds were found to be effective in scavenging DPPH radicals and tyroinase inhibition. The aim of this study was to evaluate the anti-tyrosinase and antioxidant activity of three different extracts from the ultrasound-assisted method and their metal complexes from A. ursinum to discover new candidates for food additives, cosmetic and pharmaceutical products. METHODS: Water, 70% ethanol and absolute ethanol extract of Allium ursinum and their man- ganese and zinc-complexes were characterized by FT-IR and UV-Vis spectra and their antioxidant and anti- tyrosinase activity determined using DPPH radical scavenging and mushroom tyrosinase assay. RESULTS: The antioxidant activity of the water extract was superior to other samples, while the 70% ethanol extract exhibited the highest anti-tyrosinase activity. Metal complex formation of the extracts led to a signifi- cantly lower antioxidant effect. The tyrosinase inhibition strongly related to the metal ion and extraction sol- vent. All the zinc complexes had lower anti-tyrosinase activity than their extracts, while the manganese com- plex of the water and absolute ethanol extracts exhibited higher anti-tyrosinase activity than related extracts. CONCLUSIONS: This study shows that Mn complex of A. ursinum extracts, based on the solvent extraction, could increase tyrosinase inhibition activity and could be a good candidate for intended cosmetic applications and food additives.

In Vivo Anti-Malarial Activity of Heracleum persicum Fruit Extract, in Combination with Chloroquine against Chloroquine-Sensitive Strain of Plasmodium berghei.

BACKGROUND: We evaluated the anti-malarial activity of Heracleum persicum individually and in combination with chloroquine. METHODS: This study was conducted at the School of Public Health, Tehran University of Medical Sciences, Tehran, Iran in 2015-2016. The Peter's method was used for determining fifty percent effective dose (ED50) of the H. persicum extract and chloroquine individually against chloroquine sensitive P. berghei in small white mice. Six experimental groups for H. persicum and 6 groups for chloroquine and two control group (positive and negative) were considered for determination of ED50. Interaction between H. persicum and chloroquine also was evaluated based on fixed ratios method. Ratios of 0/100, 20/80, 40/60, 60/40, 80/20, 100/0 of ED50 of chloroquine and H. persicum respectively were tested against the parasite. Then inhibitory effects of two drugs were calculated and plotted in the relevant graphs. RESULTS: Overall, 1500 mg/kg, and 1000 mg/kg concentrations of H. persicum against P. berghei resulted in ED50 and ED74 respectively. ED50 of chloroquine against the parasite was obtained as 1.4 mg/kg of mouse body weight. Moreover, combination of H. persicum and chloroquine showed a weak potentiation in ratios of 40/60 (chloroquine +H. persicum) with 64% inhibition, but not in other ratios. CONCLUSION: Although H. persicum individually showed a reasonable antimalarial efficacy against chloroquine sensitive P. berghei, in combination with chloroquine it showed additive or antagonism result except in ratios of 40%CQ+60%HP.

Validated Spectrophtometric Method for Simultaneous Determination of Buprenorphine and Naloxone in Pharmaceutical Dosage Forms.

Buprenorphine is a partial mu agonist and kappa antagonist which is used for the treatment of pain and opioid addiction. A mixture of buprenorphine hydrochloride and naloxone hydrochloride has been approved for the treatment of opioid dependence. In this study a third order derivative spectrophotometric method based on zero-crossing technique has been used for the simultaneous determination of buprenorphine hydrochloride and naloxone hydrochloride in tablets. The measurements were carried out at wavelengths of 257.8 (zero-crossing point of naloxone hydrochloride) and 252.2 nm (zero-crossing point of buprenorphice hydrochloride) for buprenorphine hydrochloride and naloxone hydrochloride, respectively in the third order derivative spectra obtained in methanol and 0.1 M NaOH (50:50) as solvent. The method was found to be linear in the range of 20-80 µg/mL for buprenorphine hydrochloride and 5-20 µg/mL for naloxone hydrochloride. The within-day and between-day coefficient of variation and error values were less than 2.5% and 1.8%, respectively. The proposed method was successfully used for simultaneous determination of these drugs in pharmaceutical dosage form without any interference from excipients or need to prior separation before analysis.

Fabrication and biological evaluation of chitosan coated hyaluronic acid-docetaxel conjugate nanoparticles in CD44(+) cancer cells.

BACKGROUND: Hyaluronic acid (HA) has been used for target-specific drug delivery because of strong affinity to CD44, a marker in which overexpressed in cancer cells and cancer stem cells. Conjugation of HA to the cytotoxic agents via active targeting can improve efficacy, biodistribution, and water solubility. To be able to benefit from passive targeting as well, a nanoparticulate system by counter ion using a polycation like chitosan may lead to a perfect delivery system. METHODS: Water soluble Hyaluronic acid-Docetaxel (HA-DTX) conjugate was prepared and used to formulate chitosan-coated HA-DTX nanoparticles by polyelectrolyte complex (PEC) method and optimized using Box-Behnken design. Biological evaluation of nanoparticles was done in CD44+ cancer cells. RESULTS AND DISCUSSION: Biological evaluation of optimized formula showed IC50 of nanoparticles for 4 T1 and MCF-7 cell lines were 45.34 µM and 354.25 µM against 233.8 µM and 625.9 µM for DTX, respectively with increased cellular uptake showed by inverted confocal microscope. CONCLUSION: Chitosan-coated HA-DTX nanoparticles were more effective against CD44+ cells than free DTX. Chitosan coated hyaluronic acid-docetaxel conjugate nanoparticles fabricated and evaluated in CD44+ cancer cells.

Fourth-order derivative spectrophotometric method for simultaneous determination of pseudoephedrine and naproxen in pharmaceutical dosage forms.

Combination dosage forms of naproxen sodium and pseudoephedrine hydrochloride are used for symptomatic treatment of cold and sinus disorders. In this study, fourth-order derivative spectrophotometric method was used for simultaneous determination of naproxen sodium and pseudoephedrine hydrochloride. The method was linear over the range of 2-28 µg/ml for pseudoephedrine hydrochloride and 4-200 µg/ml for naproxen sodium. The within-day and between-day coefficient of variation values were less than 5.8% and 2.5% for pseudoephedrine hydrochloride and naproxen sodium, respectively. The application of the proposed method for simultaneous determination of naproxen and pseudoephedrine in dosage forms was demonstrated without any special pretreatment.

A Stability Indicating HPLC Method for the Determination of Fluvoxamine in Pharmaceutical Dosage Forms.

Fluvoxamine maleate is a selective serotonin reuptake inhibitor, which is used for the treatment of different types of depressive disorders. In the present study, a stability indicating HPLC method was developed and validated for the determination of fluvoxamine maleate. The chromatographic separation was carried out using a Nova-Pak CN column and a mixture of K2HPO4 50 mM (pH 7.0) and acetonitrile (60: 40, v/v) as the mobile phase. Target compounds were detected using a UV detector set at 235 nm. The developed method was linear over the concentration range of 1-80 µg/ml with acceptable precision (CV values < 2.0%) and accuracy (error values < 1.6%). The degradation studies showed that fluvoxamine maleate is relatively unstable under acidic, basic and oxidative conditions and also when exposed to UV radiation. On the other hand, the bulk powder of fluvoxamine maleate was relatively stable when exposed to visible light or heat. The proposed method was successfully applied for the determination of active ingredient of fluvoxamine dosage form without any interference from tablet excipients.

A Stability Indicating HPLC Method for the Determination of Nitisinone in Capsules.

In this study a simple and efficient stability-indicating HPLC method with short run time was developed for the determination of nitisinone. The stress degradation of nitisinone was studied in different acidic, basic, oxidative, thermal and photolytic conditions. The chromatographic separation was achieved on a Nova-Pak C18 column using a mixture of 50 mM NaH2PO4 (pH 2.5) and acetonitrile (45:55, v/v) as mobile phase. UV detection was performed at 280 nm. Good linearity was observed over the concentration range of 0.5-50 µg/ml with r(2)>0.999. The within-day and between-day precision values were less than 2%. The proposed method could be used for the determination of nitisinone in the presence of its degradation products and also dosage form excipients for the quality control purposes.

Development of a rapid derivative spectrophotometric method for simultaneous determination of acetaminophen, diphenhydramine and pseudoephedrine in tablets.

A mixture of acetaminophen, diphenhydramine hydrochloride and pseudoephedrine hydrochloride is used for the symptomatic treatment of common cold. In this study, a derivative spectrophotometric method based on zero-crossing technique was proposed for simultaneous determination of acetaminophen, diphenhydramine hydrochloride and pseudoephedrine hydrochloride. Determination of these drugs was performed using the (1)D value of acetaminophen at 281.5 nm, (2)D value of diphenhydramine hydrochloride at 226.0 nm and (4)D value of pseudoephedrine hydrochloride at 218.0 nm. The analysis method was linear over the range of 5-50, 0.25-4, and 0.5-5 µg/mL for acetaminophen, diphenhydramine hydrochloride and pseudoephedrine hydrochloride, respectively. The within-day and between-day CV and error values for all three compounds were within an acceptable range (CV<2.2% and error<3%). The developed method was used for simultaneous determination of these drugs in pharmaceutical dosage forms and no interference from excipients was observed.

A Stability-Indicating HPLC Method for the Determination of Fingolimod in Pharmaceutical Dosage Forms.

Fingolimod is an immunosuppressive agent which is used for the prophylaxis of organ transplantation rejection or multiple sclerosis treatment. In this study, systematic forced degradation studies on fingolimod bulk powder were performed to develop a stability-indicating HPLC method. Separation of fingolimod and its degradation products was achieved on a Nova-Pak C8 column. The mobile phase was a mixture of potassium dihydrogenphosphate 50 mM (pH 3.0) and acetonitrile (45:55, v/v) at a flow rate of 1 ml/min. The proposed method was linear in the range of 0.125-20 µg mL(-1). The within-day and between-day coefficients of variation were in the range of 0.6-1.2%. The developed method was successfully applied for the determination of the fingolimod amount in pharmaceutical dosage forms.

A Stability-Indicating HPLC Method for the Determination of Memantine Hydrochloride in Dosage Forms through Derivatization with 1-Fluoro-2,4-dinitrobenzene.

Memantine is chemically a tricyclic amine and is used for Parkinson's disease and movement disorders. Although several HPLC methods with different derivatization reagents have been developed for the determination of memantine in biological fluids, there are some complications which limit the use of these methods in routine analysis of memantine in in vitro tests. We established a simple, sensitive, precise, and accurate HPLC method for the quantification of memantine in dosage forms. Pre-column derivatization of memantine was performed with 1-fluoro-2,4-dinitrobenzene and the reaction product was separated on a Nova-Pak C18 column. A mixture of acetonitrile and sodium dihydrogenphosphate (pH 2.5; 0.05 M) (70: 30, v/v) was used as the mobile phase. UV detection was performed at 360 nm. Forced degradation studies were performed on a powdered tablet sample of memantine hydro-chloride using acidic (0.1 M hydrochloric acid), basic (0.1 M sodium hydroxide), oxidative (10% hydrogen peroxide), thermal (105°C), photolytic, and humidity conditions. Good linearity (r(2)=0.999) was obtained over the range of 1-12 µg mL(-1) of memantine hydrochloride with acceptable within-day and between-day precision values in the range of 0.05-0.95%. The proposed method was used for the assay determination and dissolution rate study of memantine dosage forms with excellent specificity.

A stability indicating HPLC method for the determination of clobazam and its basic degradation product characterization.

BACKGROUND: Clobazam is used for the treatment of different types of seizure and epilepsy. The present research is undertaken to study the systematic forced degradation of clobazam and to identify its main degradation product under basic conditions. METHODS: The degradation of clobazam was studied under different conditions. Clobazam and its degradation products were separated using a Nova-Pak C18 column and a mixture of KH2PO4 50 mM (pH 8.5) and acetonitrile (50:50, v/v) as the mobile phase with UV detection at 230 nm. RESULTS: The within-day and between-day precision values in the calibration range of 0.1-20 µg/ml were within 0.5-1.5%. Clobazam was relatively stable in solid from under exposure to visible and UV light and also heat. The clobazam aqueous solution of clobazam was more labile under exposure to visible and UV light. The bulk drug was significantly degraded under exposure to 2 M HCl, 0.1 M NaOH or 3% H2O2. Using the tablet powder, higher degradation rates were observed under different stress conditions. The main degradation product of clobazam under basic condition was subsequently characterized. CONCLUSION: The developed method could be used for the determination of clobazam in the presence of its degradation products with acceptable precision and accuracy. The applicability of the proposed method was evaluated in commercial dosage forms analysis.

Validated spectrophtometric method for determination of tamsulosin in bulk and pharmaceutical dosage forms.

In this study a sensitive, simple and accurate spectrophotometric method was suggested for determination of tamsulosin in bulk powder and pharmaceutical dosage form based on the formation of an ion-pair complex between the drug and bromocresol green in a buffer solution at pH 3.5. The formed yellow color complex was extracted with chloroform and measured at 415 nm. The optimum reaction conditions such as pH, reagent amount, extracting solvent and the stoichiometry of the ion-pair complex were investigated. Under the optimized conditions, the Beer's law was obeyed in the concentration range of 1-160 g/mL with acceptable correlation coefficient (r(2) > 0.9997) and precision (CV < 3%) and accuracy (error < 2%). The proposed method was successfully used for the determination of tamsulosin in pharmaceutical capsule with nosignificant interferences of excipients.

Synthesis of 6-(2-Methoxynaphthyl)-2,3-dihydro-1,2,4-triazine-3-thione as a New Reagent for Spectrophotometric Determination of Copper.

A simple, sensitive, accurate, and green spectrophotometric method for the determination of Cu(II) using newly synthesized reagent, 6-(2-methoxynaphthyl)-2,3-dihydro-1,2,4-triazine-3-thione (MNDTT), has been developed. MNDTT was synthesized based on the acylation of methoxy naphthalene and reaction of the product with amyl nitrite, which upon reaction with thiosemicarbazide yielded 6-(2-meyhoxynaphthyl)-2,3-dihydro-1,2,4-triazine-3-thione. MNDTT produces a dark red complex with copper in methanol according to the 1 : 2 stoichiometry. Beer's law was obeyed over the concentration range of 2.5-20 µg/mL with r (2) = 0.992. The limit of detection and limit of quantification were 0.33 and 1.10 µg/mL, respectively. Within-day and between-day precision values were less than 3.68%. Finally, the method has been applied to a dental alloy (110-plus) successfully and the results were compared with atomic absorption method. The results showed that there was no significant difference between the two methods (P > 0.05).

Discovery of new angiotensin converting enzyme (ACE) inhibitors from medicinal plants to treat hypertension using an in vitro assay.

BACKGROUND AND PURPOSE OF THE STUDY: Angiotensin converting enzyme (ACE) inhibitors plays a critical role in treating hypertension. The purpose of the present investigation was to evaluate ACE inhibition activity of 50 Iranian medicinal plants using an in vitro assay. METHODS: The ACE activity was evaluated by determining the hydrolysis rate of substrate, hippuryl-L-histidyl-L-leucine (HHL), using reverse phase high performance liquid chromatography (RP-HPLC). Total phenolic content and antioxidant activity were determined by Folin-Ciocalteu colorimetric method and DPPH radical scavenging assay respectively. RESULTS: Six extracts revealed > 50% ACE inhibition activity at 330 µg/ml concentration. They were Berberis integerrima Bunge. (Berberidaceae) (88.2 ± 1.7%), Crataegus microphylla C. Koch (Rosaceae) (80.9 ± 1.3%), Nymphaea alba L. (Nymphaeaceae) (66.3 ± 1.2%), Onopordon acanthium L. (Asteraceae) (80.2 ± 2.0%), Quercus infectoria G. Olivier. (Fagaceae) (93.9 ± 2.5%) and Rubus sp. (Rosaceae) (51.3 ± 1.0%). Q. infectoria possessed the highest total phenolic content with 7410 ± 101 mg gallic acid/100 g dry plant. Antioxidant activity of Q. infectoria (IC50 value 1.7 ± 0.03 µg/ml) was more than that of BHT (IC50 value of 10.3 ± 0.15 µg/ml) and Trolox (IC50 value of 3.2 ± 0.06 µg/ml) as the positive controls. CONCLUSIONS: In this study, we introduced six medicinal plants with ACE inhibition activity. Despite the high ACE inhibition and antioxidant activity of Q. infectoria, due to its tannin content (tannins interfere in ACE activity), another plant, O. acanthium, which also had high ACE inhibition and antioxidant activity, but contained no tannin, could be utilized in further studies for isolation of active compounds.

Validating a stability indicating HPLC method for kinetic study of cetirizine degradation in acidic and oxidative conditions.

A stability indicating High-Performance Liquid Chromatography (HPLC) method was validated and used to study the degradation of cetirizine dihydrochloride in acidic and oxidative conditions. The separation was carried out on a Symmetry C18 column and a mixture of 50 mM KH2PO4 and acetonitrile (60:40 v/v, pH = 3.5) was used as the mobile phase. The method was linear over the range of 1-20 µg/mL of cetirizine dihydrochloride (r(2) > 0.999) and the within-day and between-day precision values were less than 1.5%. The results showed that cetirizine dihydrochloride was unstable in 2 M HCl and 0.5% H2O2. The kinetics of the acidic degradation showed a pseudo-first-order reaction in the temperature range of 70-90°C. In addition, the kinetics of hydrogen peroxide mediated degradation was pseudo-first-order in the temperature range of 50-80°C.

Isolation, identification and molecular docking studies of a new isolated compound, from Onopordon acanthium: a novel angiotensin converting enzyme (ACE) inhibitor.

ETHNOPHARMACOLOGICAL RELEVANCE: Onopordon acanthium (also known as Scotch thistle) is a medicinal plant of the Asteraceae family that is widely distributed in Europe and Asia. This plant has been long used in traditional medicine as a hypotensive, cardiotonic and diuretic agent. AIM OF THE STUDY: The present study is designed to isolate an active compound with ACE inhibition activity from O. acanthium, measure antioxidant activity, predict domain specificity and pharmacokinetic properties of the isolated compound. MATERIALS AND METHODS: Methanolic extract of O. acanthium seeds, has been subjected to a repeated column chromatography to give a pure compound with Angiotensin Converting Enzyme (ACE) inhibition activity. The ACE inhibition activity was determined using hippuryl-L-histidyl-L-leucine (HHL) as substrate in an in vitro ACE assay. Structure of the pure compound, isolated from O. acanthium has been established by spectroscopic methods, including Infrared (IR), Nuclear Magnetic Resonance (NMR) and Mass spectrum analysis. In addition, antioxidant activity of the new isolated compound, was measured using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay and compared with those of BHT and Trolox as positive controls. Enzyme type inhibition and ACE-C or N domain specificity of the new compound was further evaluated through molecular modeling and docking studies. RESULTS: Structure of the pure compound, isolated from O. acanthium (83±1% ACE inhibition activity at concentration of 330 µg/ml), has been established. The isolated compound possessed acceptable antioxidant activity (IC50 value of 2.6±0.04 µg/ml) in comparison with BHT (IC50 value of 10.3±0.15 µg/ml) and Trolox (IC50 value of 3.2±0.06 µg/ml). Molecular docking predicted competitive type enzyme inhibition and approximately similar affinity of the isolated compound for ACE-C and N domains. CONCLUSION: The results derived from computational and in vitro experiments, confirm the potential of the isolated compound, from O. acanthium as a new antihypertensive compound and give additional scientific support to an anecdotal use of O. acanthium in traditional medicine to treat cardiovascular disease such as hypertension.

Derivative spectrophotometric method for simultaneous determination of clindamycin phosphate and tretinoin in pharmaceutical dosage forms.

A derivative spectrophotometric method was proposed for the simultaneous determination of clindamycin and tretinoin in pharmaceutical dosage forms. The measurement was achieved using the first and second derivative signals of clindamycin at (1D) 251 nm and (2D) 239 nm and tretinoin at (1D) 364 nm and (2D) 387 nm.The proposed method showed excellent linearity at both first and second derivative order in the range of 60-1200 and 1.25-25 µg/ml for clindamycin phosphate and tretinoin respectively. The within-day and between-day precision and accuracy was in acceptable range (CV<3.81%, error<3.20%). Good agreement between the found andadded concentrations indicates successful application of the proposed method for simultaneous determination of clindamycin and tretinoin in synthetic mixtures and pharmaceutical dosage form.

A New extractive spectrophotometric method for determination of rizatriptan dosage forms using bromocresol green.

BACKGROUND AND THE PURPOSE OF THE STUDY: Rizatriptan is used effectively for the treatment of migraine headache. In this study, a simple, rapid and low cost spectrophotometric method based on the ion-pair complexation is proposed for the determination of rizatriptan in raw material and dosage forms. METHODS: The ion-pair complexation using bromocresol green as reagent was performed in a buffer solution and the absorbance was measured by a spectrophotometer. The ion-pair formation conditions were optimized and the accuracy and precision of the method were calculated. RESULTS AND MAJOR CONCLUSION: Best results were achieved by using 6 ml of the bromocresol green reagent in the presence of phosphate buffer (pH 3.0). The stoichiometry of the resulted complex was 1:1. The within-day and between-day precision values were lower than 2.9 and 1.8 percent for the calibration range of 0.5-50 and 10-100 µg/ml, respectively. The proposed method was successfully used for the determination of rizatriptan in dosage forms without any interference.