IN VITRO MEASUREMENT OF TOTAL ANTIOXIDANT CAPACITY OF CRATAEGUS MACRACANTHA LODD LEAVES.

UNLABELLED: Crataegus macracantha Lodd, family Rosaceae, is a very rare species in Europe, and unlike Crataegus monogyna is less investigated for pharmacologic activity. AIM: To analyze the ability of the lyophilisate of extract obtained from leaves of Crataegus macracantha Lodd (single plant at the Iasi Botanical Garden) to capture free radicals in vitro. MATERIAL AND METHODS: The lyophilisate obtained in Department of Pharmacognosy, Faculty of Pharmacy, "Grigore T. Popa" University of Medicine and Pharmacy Iasi. The decreased absorbance of chromophore chlorpromazine radical cation in the presence of the lyophilized solutions was studied spectrophotometrically. The indicator radical cation, obtained by oxidation of chlorpromazine by potassium persulfate, has the maximum absorbance at 525 nm. Ascorbic acid was used as a standard antioxidant. RESULTS: The absorbance of radical solution was determined after the addition of a certain amount of lyophilisate at different time intervals. The antioxidant activity was calculated using the calibration curve obtained by plotting the variation in radical solution absorbance depending on ascorbic acid concentration. For each ascorbic acid concentration the area under the curve was calculated from plotting the percentage inhibition of the absorbance at two pre-established time intervals. CONCLUSIONS: The results confirm the antioxidant activity of the leaves of Crataegus Macracantha Lodd and by optimizing the proposed analytical methods the antiradical activity can be quickly evaluated with minimal reagent consumption.

[Not Available]. / Le microdosage colorimetrique des phosphates dans des milieux divers (Produits biologiques, pharmaceutiques et alimentaires).

A colorimetric micromethod is proposed for determination of phosphates in various materials (biological substances, pharmaceuticals and food). It is based on precipitation of magnesium uranyl phosphate, which is then dissolved in dilute sulphuric acid, and the uranyl ion is determined spectrophotometrically via the dark red colour of uranyl ferrocyanide. The sensitivity is 0.01 mg of P, and the average error 1%. The method is very simple and applicable to many types of sample.

[Not Available]. / Nouvelle microcolorimétrie des nitrites.

A micromethod for determination of nitrite is based on a reaction with o-tolidine, forming a yellow-orange product suitable for spectrophotometry. The sensitivity is 0.05 mug of nitrite ml . The method is suitable for analysis of potable and mineral waters, and has a mean relative error of 1%.

Phosphomolybdic acid as a reagent for the spectrophotometric determination of certain phenothiazine derivatives of pharmaceutical products.

Phosphomolybdic acid is used as reagent for colorimetric determination of phenothiazine derivatives in various pharmaceutical products. The reagent oxidizes the derivative to a cationic free radical, with which it then forms a coloured salt. The method is simple and rapid. A preliminary extraction is necessary if certain reductants (sulphite or ascorbic acid) are present as stabilizers of the pharmaceuticals, as otherwise they reduce the reagent to phosphomolybdenum blue.

Schiff bis bases: analytical reagents. II. Spectrophotometric determination of manganese from pharmaceutical forms.

By condensing ethyl-o-hydroxybenzene with ethylene diamine, and 1-ethyl-salicylidene bis ethylene diamine, a Salen-type Schiff bis base is obtained. These Schiff bis bases present a good capacity of complexing the Mn(II) ions, resulting brown complexes. In this paper, the results of a study concerning the use of the Schiff bis base as reagent in spectrophotometric determination of the Mn(II) is presented. The above mentioned Schiff bis base forms a brown complex with Mn(II) cation, with maximum absorbance at 460 nm, and molar absorbtivity (epsilon)=9.8 x 10(4). The complex with Mn(II) presents a maximum stability at pH 6.0. The combination ratio was established by isomolar series method, and it is 1:2 (metal:ligand). The calculated apparent stability constant is beta(n)=2.943 x 10(-5). The absorbance is proportional to Mn(II) concentration in the range of 10-70 microg ml(-1). In this range, the Lambert-Beer law is respected, the linearity coefficient being 0.9989, S.D.=0.83, R.S.D.=0.88 (n=7). In these conditions, the complexation reaction of Mn(II) is interfered by other cations, Fe(II); Fe(III); Ni(II). The results obtained for spectrophotometric determination of Mn(II) using this Schiff base as reagent were successfully applied to pharmaceutical products containing Mn(II) cation.

Cyclosporine bioavailability of two physically different oral formulations.

To assess the comparative bioavailability of two cyclosporine capsule products with different pharmaceutical formulation an open randomized two-period cross over study was conducted in 24 healthy volunteers. Our results, obtained from cyclosporine HPLC determination onto the whole blood samples collected, show that the test cyclosporine non-SMEDDS formulation was not bioequivalent cyclosporine SMEDDS formulation due to a statistically significantly lower absorption rate. The outcome does not support free and full interchangeability in chronic stable graft recipients of the two products studied, unless validated clinical and laboratory conversion protocols for each kind of organ transplantation are enforce.

Possible involvement of the L-arginine-nitric oxide pathway in the modulation of stress-induced analgesia.

The possible participation of nitric oxide (NO) in pain modulation and stress analgesia was studied in adult Wistar rats. Cerebral citruline as a coproduct of NO from L-arginine increased from the mean value 5.6 +/- 0.4 nM/mg.Pt. to 8.9 +/- 0.5 nM/mg.Pt. in acute restraint stress. In high doses (50 mg/kg body weight), intraperitoneal administration of L-arginine caused a small and transient decrease of the tail-flick latencies to the thermal stimulus, without significant changes of the stress analgesia induced by restraint stress. In animals pretreated with N-nitro-L-arginine methyl ester (NAME) a progressive increase of the latency time was obtained and the increased latencies induced by acute immobilization appeared significantly potentiated. These results offer new indirect evidence in favour of the modulatory role of NO in thermoalgesic sensitivity and stress-induced analgesia.

Evaluation of free radical scavenging activity of some antioxidants.

UNLABELLED: Measurement of free radical scavenging capacity of antioxidants is influenced by a number of parameters that depend on reaction rate. AIM: A new method for assessing the free radical scavenging activity in which the influence of antioxidant concentration and reaction rate on chlorpromazine radical cation absorbance are simultaneously monitored. MATERIALS AND METHODS: The change in radical solution absorbance at 525 nm within a fixed time at different concentrations of the standard antioxidant--ascorbic acid (AA) are determined and percent inhibition is calculated. RESULTS: The percent inhibition of color was plotted versus time, and the area under the curve was calculated for each concentration of the standard antioxidant. The calibration curve was obtained by plotting the area under the curve versus ascorbic acid concentration. The antioxidant activity of the samples was calculated using the regression line equation (r2 = 0.9991) and expressed as ascorbic acid molar equivalents (AAE) depending on the unit of measurement chosen for the tested product. CONCLUSIONS: The proposed method takes into account the two parameters influencing the kinetics of the reaction between antioxidant and radicals, namely the antioxidant concentration and fixed time for measuring absorbance.

Spectrophotometric method for estimation of bisoprolol fumarate in tablets.

UNLABELLED: Bisoprolol fumarate is prescribed for the treatment of hypertension and angina pectoris. AIM: The purpose of this study was to develop a simple, sensitive, accurate, and reproducible method for estimation of bisoprolol fumarate in tablets. MATERIAL AND METHODS: The proposed method was based on a yellow colored complex formed with tropaeolin 00, extractable in dichloromethane with maximum absorbance at 412 nm. The method was validated statistically. RESULTS: The linearity domain was observed in the concentration of 5-30 microg/ml. The recovery studies confirmed the accuracy of the proposed method. CONCLUSIONS: The proposed method can be applied for the routine analysis of bisoprolol from formulations.

Spectrometric determination of total antioxidant activity in chlorpromazine radical cation - ascorbic acid system.

UNLABELLED: Through univalent oxidation chlorpromazine forms a colored and relatively stable radical cation with maximum absorbance at 525 nm, considered a redox mediator in a number of b iochemical reactions. AIM: To develop a spectrometric method for the determination of total antioxidant activity based on the reaction of chlorpromazine radical capture by ascorbic acid (standard antioxidant). MATERIAL AND METHODS: The calibration curve was drawn by monitoring the decrease in the absorbance of the preformed radical solution (obtained by oxidation of chlorpromazine by potassium persulfate in an acidic environment) depending on ascorbic acid concentration. RESULTS: The method was validated. In the ascorbic acid concentration range 10-100 microM/L linearity was good (r2 = 0.9991). Limit of detection (LOD) was 3.13 microM/L and limit of quantification (LOQ) was 9.49 microM/L. System precision (RSD = 0.75%), method precision (RSD = 4.50%) and intermediate precision (RSD = 4.63%) were determined. The average recovery of 101.7% for the concentration range 91.1 -105.9% confirmed the accuracy of the method. CONCLUSIONS: The method has a good linearity, precision, accuracy, and is easy to use for evaluation of antioxidant action of different products. Total antioxidant activity is expressed as ascorbic acid molar equivalents (AAE). The method has the advantage of using a radical involved in redox processes in the body.

Spectrophotometric determination of enalapril through ternary complex formation.

UNLABELLED: Enalapril is an angiotensin-converting enzyme inhibitor widely used in the treatment of hypertension and heart failure. AIM: To design and validate a simple and sensitive spectrophotometric method. This method is based on a ternary complex formation between enalapril, copper (II), and bromothymol blue, in a slightly alkaline environment, complex that is extractable in chloroform, with an absorption maximum at 426 nm. MATERIALS AND METHODS: To a volume of 1 milliliter of standard enalapril drug solution in concentration range 200/500 microg/ml, two milliliters of Cu (II) chloride solution 0.2 % followed by 2 ml bromothymol blue solution 0.01% and 0.5 ml of pH 8 buffer solution (Britton-Robinson) were added. The complex was two times extracted with 2 x 1.5 ml chloroform. Ten minutes after extraction, absorbance was measured at 426 nm against blank. RESULTS: Our spectrophotometric method was validated by determining linearity, limit of detection and quantification, system and method precision, and method accuracy. The method showed a good linearity in the range 200/500 microg/ml (correlation coefficient r = 0.9993). Limit of detection (LD) was 9.907 microg/ml and limit of quantification (LQ) 33.024 microg/ml. Precision showed an RSD = 1.83%, and accuracy with mean recovery is 102.6% in the range 98.8/103.9. CONCLUSIONS: The ternary complex formed under the above-mentioned condition and measured spectrophotometrically can be regarded as an ion-association complex between the metal-drug cation and bromothymol blue anion. The experimental results demonstrated a good sensitivity.

[Development and validation of a GC-MS method for determination of nicotine in tobacco]. / Dezvoltarea si validarea unei metode GC-MS pentru determinarea nicotinei din tutun.

UNLABELLED: DETERMINATION FROM TOBA gas chromatographic-mass spectrometric method (GC/MS) for determination of nicotine from tobacco products was developed and validated. MATERIAL AND METHOD: It was used an Agilent Technologies 7890A Chromatograph equipped with a DB 5 MS column (30 m x 0.25 mm, 0.25 microm) and Agilent Technologies 5975C inert MSD detector. The temperature program began from 100 degrees C and increase with 10 degrees C/min to 190 degrees C than with 20 degrees C/min to 280 degrees C, constant for 5 min, with He as carrier gas (1 ml/min) and MS detection. The temperature of the source was 230 degrees C and the temperature of the quadrupole was 150 degrees C. The parameters of validation were studied according to the international requirements. RESULTS: Finally, the method established has following parameters: is linear in the 1.01-201.8 microg/ml range, has the detection limit of 3.6 microg/ml, the quantification limit of 10.8 microg/ml, is precise (RSD < 5%, n = 9) and accurate (mean recovery is 100.28% in 99.08-100.96 microg/ml range). CONCLUSION: The method can be used to determine nicotine from tobacco products, with good results.

Spectrometric method for the assay of ramipril using molybdophosphoric acid.

UNLABELLED: Ramipril is a drug of the angiotensin converting enzyme inhibitor class. AIM: A new molecular absorbance spectrometric method was developed for the assay of ramipril using molybdophosphoric acid in acidic medium. MATERIAL AND METHODS: The reaction product showed a maximum absorbance at 361 nm. The optimum conditions of the reaction were established. The developed method was validated. RESULTS: The method showed a good linearity in the range of 8 - 36 microg/ml (correlation coefficient r = 0.9996). The detection limit (LD) was 0.86microg/ml and the quantification limit (LQ) 2.88 pg/ml. Precision and accuracy were determined (RSD = 1.15%); mean recovery was 98.90% in the 97.13-101.03% concentration range. CONCLUSIONS: The proposed method is simple, easy to perform, sensitive, linear, precise, accurate and robust.

[Validation of a HPLC method for ochratoxin A determination]. / Validarea unei metode HPLC pentru determinarea ochratoxinei A.

Ochratoxin A is a mycotoxin produced by various species of Aspergillus and Penicillium. Ochratoxin A has been detected in cereals and cereal products, coffee beans, beer, wine, spices, pig's kidney and cow's milk. For ochratoxin A, a HPLC method was developed and validated. Ochratoxin A was determined by RP-HPLC, using a liquid chromatograph type HP 1090 Series II, equiped with a fluorescence detector. The analysis was performed with a Phenomenex column, type Luna C18(2) 100A (150 x 4.6 mm; 5 microm) with a mobile phase consisting of a mixture of acetonitrile/water/acid acetic (99/99/2), a flow of 0.7 mL/min. For detection, the wavelenght of excitation was 228 nm and wavelenght of emision was 423 nm. The calibration graph was linear in 6.25-50 ng/mL concentration range (r2 = 0,9991). The detection limits was 1.6 ng/mL and the quantification limit was 4.9 ng/mL. The method precision (RSD = 2.4975%) and the accuracy (recovery was 100.1%) were studied. The HPLC method was applyed for ochratoxin A from food samples with good results.

[Contribution to the mobile phase selection for HPLC separation in order to make the identification and quantitative determination using the UV-VIS detector]. / Contributii la alegerea fazei mobile in vederea separarii prin HPLC si identificarea--determinarea cantitativa a unor antiinflamatoare nesteroidiene folosind detectorul UV-VIS.

UNLABELLED: The aim of this study is to elaborate a HPLC with UV detection method for the analysis of some AINS, method which can be transferred in a system with mass spectrometry detection. MATERIAL AND METHODS: More AINS drugs (diclofenac sodium, ibuprophen, ketoprofen, meloxicame, piroxicame, tenoxicame, nimesulide, phenylbutazone and indomethacin) were studied by UV spectrophotometry and by HPLC. RESULTS: The UV spectra of the studied AINS were recorded in pure solvents or in a mixture of solvents in order to establish the absorbance that will be used for HPLC detection in UV. To establish the optimum chromatographic parameters, the influence of the flow and of the composition of the mobile phase (both of the aqueous and organic phases) were studied too. CONCLUSIONS: The proposed procedure is easy and relatively with low cost regarding the MS detection.

The study of aminophenazone radical cation and its interaction with some antioxidants.

UNLABELLED: The purpose of this study was to identify both the possibilities of in vitro generation of 1-phenyl-2,3-dimethyl-4-dimethylamino-5-pyrazolone (aminophenazone) radical cation (abbreviated APH*+) and the mechanisms of interaction with different antioxidants. MATERIAL AND METHOD: The chromophore radical cation APH*+ was obtained by oxidizing APH with ammonium persulfate. The stages of the oxidation reaction and the production of the intermediate compounds were studied by spectrophotometrical and potentiometric titration. As standard antioxidant there was used a catechin solution in distilled water with concentrations ranging between 50-250 microg/mL, in weak alkaline environment. RESULTS: The oxidation of APH with ammonium persulfate developed in four stages, involving the production of some intermediate compounds with different colorations. A mechanism for this reaction has been proposed. The radical cation forms to an APH - ammonium persulfate molar ratio of 2 : 1. In the presence of an antioxidant (catechin) the radical cation APH*+ forms to a molar ratio APH-ammonium persulfate of 1:1, in weak alkaline environment. It has an absorbtion maxima at lambda = 540 nm. The molar extinction coefficient of APH*+ at lambda = 540 nm has been calculated. Its absorbance was proportional with catechin concentration in the range 50-250 microg/mL (r2 = 0.9988). CONCLUSION: The APH-ammonium persulfate system can be used for spectrophotometric determination of total polyphenols in different systems.

Measurement of total antioxidant activity with chlorpromazine radical cation.

UNLABELLED: In the present study there is described a simple and sensitive method for the evaluation of antioxidant activity in chlorpromazine-Fe (III) system using ascorbic acid as standard. MATERIAL AND METHOD: Chlorpromazine [2-chloro-N-(3-dimethyl-aminopropyl)-phenothiazine] is oxidized in acidic media by Fe (III) with the formation of a stable radical cation characterized by an intense visible absorbtion band (lambda = 540 nm). The optimum parameters for the stability of the radical cation have been studied: molar ratio chlorpromazine-Fe (III), pH, solvents. Spectrophotometric methods have been used in these studies. RESULTS: The chlorpromazine radical cation is stable in the acidic media (pH = 3) at a molar ratio chlorpromazine: Fe (III) of 2: 1. Ascorbic acid reduces these radicals and induces a decrease of absorbance. Percent inhibition was calculated and plotted as a function of the concentration of standard antioxidant solutions. The results show that percent inhibition varies in a linear manner with the ascorbic acid concentration. Percent inhibition is higher when the antioxidant solution is added after generation of radical cation. CONCLUSIONS: It has been developed a method for evaluating antioxidant activity in the chlorpromazine: Fe (III) system using ascorbic acid as a standard. The method is fast, simple and sensitive; it can be applied for the detection and evaluation of the antioxidant activity of simple or complex systems.

[Turbidimetric determination of tenoxicam using molybdophosphoric acid]. / Determinarea turbididimetrica a tenoxicamului folosind ca reactiv solutia de acid fosfomolibdenic.

UNLABELLED: A turbidimetric method was developed for tenoxicam quantification using the complexing reaction with molybdophosphoric acid in hydrochloric acid medium; the complex has a maximum of absorbance at 369 nm. MATERIALS AND METHOD: The practical working conditions were established. In the 2.5 division by 15.0 microg/mL concentration range of tenoxicam in 1N hydrochloric acid, were added 1 mL of 1% molybdophosphoric acid solution and 1 mL of 0.1% sodium lauryl sulphate. The stability of product was evaluated over 60 minutes. The developed method was validated. RESULTS: The method showed a good linearity in the range of 2.5 division by 15.0 microg/mL (the correlation coefficient r = 0.9996). The detection limit (LD) was 0.44 microg/mL and the quantification limit (LQ) was 1.47 microg/mL. There were established the precision (RSD = 1.82%) and the accuracy (mean recovery is 100.79% in 97.55 division by BY 04.41% the range). CONCLUSIONS: The experimental results demonstrated a good sensibility. The specific absorptivity for this method is A(1cm,369nm)(1%) higher than tenoxicam in hydrochloric acid medium A(1cm,354nm)(1%).

[Construction and characterisation of a selective membrane electrode for the assay of lisinopril]. / Constructia si caracterizarea unui electrod membrana selectiv pentru determinarea lisinoprilului.

UNLABELLED: In order to analyze lisinopril using a selective membrane electrode, the electro-active compound was obtained and included in the selective membrane. The new analysis method was developed and validated. MATERIAL AND METHOD: The electro-active compound was obtained through the precipitation of lisinopril in acidic media using silicowolframic acid solution. The membrane was obtained by mixing the electro-active compound with polyvinylchloride using tetrahydrofurane as solvent. The solution was evaporated in order to obtain a thick membrane, which was then attached at one end of a polyvinylchloride tube. The internal Ag/AgCl reference electrode was inserted in that tube. The assembly was filled with a lisinopril internal solution. The electrode was characterized by establishing the following characteristics: electrode slope, selectivity, optimal pH range, response time and life-time period. The developed method was validated. RESULTS: The method showed a good linearity in the range of 10(-7) and 10(-2)M (the correlation coefficient r = 0.9991). The detection limit (LD) was 8,66 x 10(-8)M and the quantification limit (LQ) was 7,8 x 10(-7)M. There were established the precision (RSD = 1.73%) and the accuracy (mean recovery was 99.92%) CONCLUSIONS: The experimental results demonstrated a good sensibility.

Evaluation of some pharmaceutical formulations of lisinopril through dissolution testing.

AIM: Lisinopril is a drug of the angiotensin converting enzyme (ACE) inhibitor class that is primarily used in treatment of hypertension, congestive heart failure, heart attacks and also in preventing renal and retinal complications of diabetes. We compared the dissolution profiles of Lisinopril 20 mg tablets (Antibiotice S.A. lasi) and Zestril 20 mg tablets (Astra Zeneca). MATERIAL AND METHOD: Because lisinopril is a third class active substance, we performed dissolution tests in standard media at three pH values: 1.2, 4.5 and 6.8 using the paddle apparatus at 75 rpm. RESULTS: Both pharmaceutical formulations present a dissolution percentage more than 85% (Q) of the labeled amount. CONCLUSION: Both pharmaceutical formulations present similar dissolution profile. Key words: