An innovative validated spectrofluorimetric method for determination of Lisinopril in presence of hydrochlorothiazide; application to content uniformity testing.

A new sensitive and discriminating spectrofluorimetric method has been developed and validated for determination of Lisinopril, one of the angiotensin converting enzyme inhibitors, in its pure bulk form and pharmaceutical tablets. The reaction of Lisinopril with ethylacetoacetate and formaldehyde in acidic buffered medium (pH3.8) has yielded a pale yellow product that exhibited a high fluorescence measured at 438nm after excitation at 350nm. All the experimental parameters affecting the formation and stability of the produced fluorophore were carefully investigated and optimized to give the maximum sensitivity. The fluorescence intensity was directly proportional to the drug concentration in the range of 0.5-4.5µg/mL with a limit of detection equal to 0.16µg/mL. The method was successfully applied in the analysis of the commercially available pharmaceutical tablets containing the single drug or its binary mixtures with Hydrochlorothiazide. Furthermore, the developed procedure was adapted for studying the content uniformity test of some dosage forms containing the cited drug.

Selective spectrofluorimetric method for determination of Lisinopril in pharmaceutical preparations and in presence of hydrochlorothiazide: Application to content uniformity testing.

A novel sensitive and cost-effective spectrofluorimetric method has been developed and validated for determination of lisinopril (an angiotensin converting enzyme inhibitor) in its pure form and pharmaceutical preparations. The method is based on the reaction of the drug with ninhydrin and phenylacetaldehyde in buffered medium (pH 7.0) to form a highly fluorescent product measured at 460 nm after excitation at 390 nm. Different experimental parameters were optimized and calibration curve was constructed. The fluorescence-concentration relationship was linear in the range of 0.15-4.0 µg mL-1 . The calculated Limit of detection (LOD) and Limit of quantitation (LOQ) were 0.04 and 0.12 µg mL-1 , respectively. The method was successfully applied for the analysis of pharmaceutical preparations containing the studied drug either alone or co-formulated with hydrochlorothiazide. The obtained results were in agreement with those of the reported method in respect to accuracy and precession. Moreover, the method was applied content uniformity testing according to United States Pharmacopeia (USP) guidelines.

Simultaneous determination of ACE inhibitors and dexibuprofen in active pharmaceutical ingredient, formulations and human serum by RP-HPLC.

The contemporary work describes a rapid and cost effective reversed phase High Performance Liquid Chromatography (RP-HPLC) method for the quantification of Captopril, Lisinopril and Dexibuprofen (DXP) simultaneously in dosage formulations, active pharmaceutical ingredients and human serum. The chromatographic system included LC-20A pump, Sil-20A auto sampler and SPD-20A UV/visible detector. The estimation was carried out by using a C18 (5µm, 250 ×4.6 mm) column with mobile phase methanol: water (80:20 v/v, pH 3.0) at 230 nm with a flow rate of 1.0 ml•min-1. The retention time of Dexibuprofen was 5.4 min while that of Captopril and Lisinopril were found to be 3.2 and 1.8 minutes respectively. There was no considerable variation exists in between the tested drug spiked in serum and the extent recovered, without interference of serum in concurrent approximation. The method developed was found to be precise, selective and validated for precision, linearity, specificity, accuracy, limit of detection and limit of quantitation. There is no such method reported earlier for the determination of ACE Inhibitors and DXP simultaneously. The present study helps in assessing the co-administration of both drugs in treatment and can be employed for quality control analysis and drug-drug interaction studies.

Application of quantitative NMR for purity determination of standard ACE inhibitors.

This study investigated the accuracy of the quantitative NMR method for purity determination of ACE inhibitors reference standards and the discovery of two pairs of new diastereoisomers. Six types of ACE inhibitors, imidapril hydrochloride, benazepril hydrochloride, lisinopril, enalapril maleate, quinapril hydrochloride, and captopril were quantificated and validated for the qNMR method by discussing factors that affect parameters of the qNMR experiment, internal standards, integration, pH-effect, and uncertainty. The results were compared with data obtained by the mass balance method. The study found that maleic acid influenced the quantification of captopril in deuteroxide because of a chemical reaction. The mixtures of the reaction products were isolated by HPLC and structurally elucidated by NMR as two pairs of new diastereoisomers, 1-[(2S,4R)-thio-2-methylpropionyl-5-d-ethanedicarboxylicacid]-L-proline and 1-[(2S,4S)-thio-2-methylpropionyl-5-d-ethanedicarboxylicacid]-L-proline. The results showed that the accuracy and precision of quantitative (1)H NMR spectroscopy satisfied the requirements for quantitative analysis of chemical reference standards and provided a simple, rapid, and reliable method for purity determination of ACE inhibitors systematically.

Stability-indicating LC method for the simultaneous determination of lisinopril and hydrochlorothiazide.

A simple and rapid stability-indicating liquid chromatographic method was developed and validated for the simultaneous determination of lisinopril and hydrochlorotiazide (HCTZ) in drug substances and dosage forms in the presence of degradation products. Forced degradation studies were conducted on the pure drugs under hydrolytic, oxidative, thermal and photolytic conditions. A chromatographic separation of the two drugs and its degradation products was achieved with an RP-18 column, using methanol, acetonitrile and phosphate buffer (pH 7.1; 0.05 M) (15:15:70, v/v/v) as mobile phase at a flow rate of 0.8 mL min(-1) and UV detection at 210 nm. Lisinopril and HCTZ were well resolved from its degradation products showing the stability-indicating capability of the method. The described method was linear over a range of 40-200 µg mL(-1) for lisinopril and 25-175 µg mL(-1) for HCTZ. The assay was also selective, accurate and precise for lisinopril and HCTZ determination. This method represents an alternative to the United States Pharmacopeia (USP) method showing shorter retention time. The method was successfully applied for determination of lisinopril and HCTZ in combined commercial tablets. The results showed that the proposed method was found to be suitable for quantitative determination and the stability study of lisinopril and HCTZ in pharmaceutical samples.

Analysis of purity profiles of generic lisinopril tablets marketed in Croatia.

In view of an increasing number of generic drugs emerging, a comparative study was performed including the approved lisinopril preparations in the form of tablets marketed in Croatia, to compare purity profiles of generic drugs versus the original medicinal product. Several batches of each individual medicinal product at different stages of their shelf life were analyzed. Impurities were determined by means of high performance liquid chromatography (HPLC). Impurity profiles were demonstrated to be specific for each individual drug. Original drug, as compared to its generic copies, had the lowest values and also the lowest variability of all the tested parameters--type, total number and content of impurities--suggesting that its manufacturing process is to certain degree better controlled compared to other manufacturers. A characteristic impurity C appearing in all the assessed preparations has the lowest levels in the original drug, whereas the amount of the highest unknown impurity does not exceed 0.10% in any of the analyzed preparations. Although the original drug stands out from all the generic preparations with its purity, it can be generally concluded that, as regarding impurities levels, all the analyzed medicinal products are within the ranges of specification limits; accordingly, it is therefore not expected that, in case of lisinopril tablets, administration of the original drug as compared to any of its generic drugs, will be safer for the patient.

Purification of pharmaceutical preparations using thin-layer chromatography to obtain mass spectra with Direct Analysis in Real Time and accurate mass spectrometry.

Forensic analysis of pharmaceutical preparations requires a comparative analysis with a standard of the suspected drug in order to identify the active ingredient. Purchasing analytical standards can be expensive or unattainable from the drug manufacturers. Direct Analysis in Real Time (DART™) is a novel, ambient ionization technique, typically coupled with a JEOL AccuTOF™ (accurate mass) mass spectrometer. While a fast and easy technique to perform, a drawback of using DART™ is the lack of component separation of mixtures prior to ionization. Various in-house pharmaceutical preparations were purified using thin-layer chromatography (TLC) and mass spectra were subsequently obtained using the AccuTOF™- DART™ technique. Utilizing TLC prior to sample introduction provides a simple, low-cost solution to acquiring mass spectra of the purified preparation. Each spectrum was compared against an in-house molecular formula list to confirm the accurate mass elemental compositions. Spectra of purified ingredients of known pharmaceuticals were added to an in-house library for use as comparators for casework samples. Resolving isomers from one another can be accomplished using collision-induced dissociation after ionization. Challenges arose when the pharmaceutical preparation required an optimized TLC solvent to achieve proper separation and purity of the standard. Purified spectra were obtained for 91 preparations and included in an in-house drug standard library. Primary standards would only need to be purchased when pharmaceutical preparations not previously encountered are submitted for comparative analysis. TLC prior to DART™ analysis demonstrates a time efficient and cost saving technique for the forensic drug analysis community. Copyright © 2011 John Wiley & Sons, Ltd.

Flow-injection chemiluminescence determination of lisinopril using luminol-KMnO4 reaction catalyzed by silver nanoparticles.

A simple, sensitive, and rapid chemiluminescence method combined with flow-injection analysis was developed for the determination of lisinopril. It was found that the chemiluminescence (CL) signal arising from the reaction of alkaline luminol with KMnO(4) could be significantly increased by addition of lisinopril in the presence of silver nanoparticles. The experimental parameters affecting the chemiluminescence reaction were carefully studied and the chemiluminescence reaction mechanism was briefly discussed. Under the selected conditions, the chemiluminescence intensity was proportional to the concentration of lisinopril ranging from 0.1 mg/L to 20.0 mg/L. The detection limit was 0.027 mg/L lisinopril and the relative standard deviation for 4.0 mg/L lisinopril solution was 1.9% over eleven repeated measurements. The proposed method was applied to the determination of lisinopril in tablets and spiked human urine samples.

A novel analytical approach for reducing the consumption of organic solvents in the charge transfer-based spectrophotometric analysis: application in the analysis of certain antihypertensive drugs.

The present study describes the development of a novel analytical approach that can reduce by 50-fold the consumption of organic solvents in the charge transfer (CT)-based spectrophotometric analysis. The proposed approach employed 96-microwell assay plates for carrying out the reaction. The CT reaction between the electron-donating analyte and electron-accepting reagent was performed in microwells (200-µL of organic solvent) and the color signals were measured with a microwell-plate reader. Optimum conditions for the proposed approach were established for two antihypertensive drugs, namely ramipril (RML) and lisinopril (LSL) as model compounds for the electron-donating analytes, and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) as a π-electron acceptor. Under the optimum conditions, Beer's law was obeyed in the concentration range of 6-100 and 6-60 µg mL-1 for RML and LSL, respectively. The limits of detection were 0.97 and 1.10 µg mL-1 for RML and LSL, respectively. The precision of the methods was satisfactory; the values of relative standard deviations did not exceed 1.1 %. The proposed approach was successfully applied to the analysis of pharmaceutical dosage forms with good accuracy and precision. The results were comparable with those of the reported methods. The approach described herein is of great practical value in pharmaceutical analysis because it reduces the exposure of analysts to the toxic effects of organic solvents, lowers the analysis cost by 50-fold, and it has a high throughput property. Although the approach was validated for RML and LSL, the same methodology could be used for any electron-donating analyte for which a CT-reaction can be performed.

Lisinopril and lisinopril/hydrochlorothiazide preparations on the Belgian market: a comparative study.

Preparations containing lisinopril and the combination lisinopril/hydrochlorothiazide, and formulated as tablets were evaluated with different tests, including in vitro dissolution, assay and content uniformity, and determination of related compounds. The analytical methods were previously validated according to international guidelines. All examined products complied with the postulated requirements.

Polypill for the treatment of cardiovascular diseases part 2. LC-MS/TOF characterization of interaction/degradation products of atenolol/lisinopril and aspirin, and mechanisms of formation thereof.

A polypill for cardiovascular diseases (CVD) is under development. It is proposed to contain a combination of antithrombotic agent (aspirin), low-dose blood pressure lowering agents, i.e., angiotensin-converting enzyme inhibitor (lisinopril), one among a beta-blocker (atenolol) or diuretic (hydrochlorothiazide), and a statin (simvastatin/atorvastatin/pravatsatin, etc.). Due to the presence of multiple drugs in the same formulation, there is a strong likelihood of interaction among the drugs and/or their products. In a previous study, we observed formation of a number of interaction/degradation products from atenolol and lisinopril in the presence of aspirin. Accordingly, the purpose of this study was to characterize the resolved products using high resolution mass spectrometric and fragmentation analyses using a LC-MS/TOF system. Initially, studies were carried out on the drugs (atenolol, lisinopril and aspirin) to establish their complete fragmentation pattern. These studies were then extended to degraded samples to postulate the structures of interaction/degradation products. The characterized structures were justified through mechanistic explanations.

Characterization of impurities in the bulk drug lisinopril by liquid chromatography/ion trap spectrometry.

Two trace impurities in the bulk drug lisinopril were detected by means of high-performance liquid chromatography coupled with mass spectrometry (HPLC/MS) with a simple and sensitive method suitable for HPLC/MSn analysis. The fragmentation behavior of lisinopril and the impurities was investigated, and two unknown impurities were elucidated as 2-(6-amino-1-(1-carboxyethylamino)-1-oxohexan-2-ylamino)-4-phenylbutanoic acid and 6-amino-2-(1-carboxy-3-phenylpro-pylamino)-hexanoic acid on the basis of the multi-stage mass spectrometry and exact mass evidence. The proposed structures of the two unknown impurities were further confirmed by nuclear magnetic resonance (NMR) experiments after preparative isolation.

LC and LC-MS methods for the investigation of polypills for the treatment of cardiovascular diseases. Part 1. Separation of active components and classification of their interaction/degradation products.

"Polypill" is a fixed-dose combination (FDC) containing three or more drugs in a single pill. The same is under development for the treatment and prevention of cardiovascular diseases. In the present study, gradient LC methods were developed for simultaneous determination of the possible components of a polypill, i.e., lisinopril, aspirin and one each among atenolol/hydrochlorothiazide and atorvastatin/simvastatin/pravastatin, in the presence of a total of 13 major interaction/degradation products. The drugs and the products were well separated using a reversed-phase (C-8) column and a mobile phase comprising of acetonitrile: phosphate buffer (pH 2.3). Other HPLC parameters were flow rate, 1 ml/min; detection wavelength, 210 nm; column oven temperature, 60 degrees C; and injection volume, 5 microl. The methods were validated for linearity, precision, accuracy, and specificity. These were further modified to make them compatible for LC-MS studies by removal of the phosphate buffer and adjustment of pH by formic acid. The suitability of the methods for LC-MS studies was established by matching the theoretical mass values of the drugs with those obtained experimentally. These methods were used to determine mass values of the major interaction/degradation products, which helped to know the source of their origin.

Identification of a new impurity in lisinopril.

LC-UV scan of lisinopril revealed the presence of an unknown impurity (approximately 0.14%) at a relative retention time of 3.26 employing phosphate buffer-acetonitrile as binary gradient system while LC-MS analysis with binary gradient system comprising of a ammonia-ammonium acetate buffer (pH 5.0) and acetonitrile indicated it to be C31H41N3O7. The impurity was isolated by preparative HPLC utilizing a linear gradient of water and acetonitrile. The structural analysis of the isolated product by 1H, 13C NMR, mass spectroscopy and FT-IR revealed it to be a 4-phenyl butanoic acid derivative (CL) of lisinopril.

Characterization of a novel impurity in bulk drug of lisinopril by multidimensional NMR technique.

During the routine impurity profile of lisinopril bulk drug by HPLC (high-performance liquid chromatography), a potential impurity was detected. Using multidimensional NMR (nuclear magnetic resonance) technique, the trace-level impurity was unambiguously identified to be 2-(-2-oxo-azocan-3-ylamino)-4-phenyl-butyric acid after isolation from lisinopril bulk drug by semi-preparative HPLC. Formation of the impurity was also discussed. To our knowledge, this is a novel impurity and not reported elsewhere.

Application of pi-acceptors to the spectrophotometric determination of lisinopril in commercial dosage forms.

Two simple, rapid and sensitive spectrophotometric methods have been proposed for the determination of lisinopril in pure form and pharmaceutical formulations. The methods are based on the charge transfer complexation reaction of the drug with 7,7,8,8,tetracyanoquinodimethane (TCNQ) and p-chloranilic acid (pCA) in polar media. The lisinopril-TCNQ and lisinopril-pCA charge transfer complexes dissociate in acetone and methanol, respectively, and yield coloured TCNQ and pCA radical anions which are measured spectrophotometrically at 743 and 525 nm. Under optimised experimental conditions, Beer's law is obeyed in the concentration range of 2-26 and 25-300 microg ml-1 with molar absorptivity of 1.432x10(4) and 1.192x10(4) l mol-1 cm-1 for TCNQ and pCA methods, respectively. Both the methods have been applied to the determination of lisinopril in pharmaceutical dosage forms. Results of analysis are validated statistically.

Development and validation of a stability indicating HPLC method for determination of lisinopril, lisinopril degradation product and parabens in the lisinopril extemporaneous formulation.

The purpose of the research described herein was to develop and validate a stability-indicating HPLC method for lisinopril, lisinopril degradation product (DKP), methyl paraben and propyl paraben in a lisinopril extemporaneous formulation. The method developed in this report is selective for the components listed above, in the presence of the complex and chromatographically rich matrix presented by the Bicitra and Ora-Sweet SF formulation diluents. The method was also shown to have adequate sensitivity with a detection limit of 0.0075 microg/mL (0.03% of lisinopril method concentration). The validation elements investigated showed that the method has acceptable specificity, recovery, linearity, solution stability, and method precision. Acceptable robustness indicates that the assay method remains unaffected by small but deliberate variations, which are described in ICH Q2A and Q2B guidelines.

Synthesis and evaluation of molecularly imprinted polymers for enalapril and lisinopril, two synthetic peptide anti-hypertensive drugs.

Molecularly imprinted polymers (MIPs) for the recognition of enalapril and lisinopril were prepared using 4-vinylpyridine as the functional monomer. Following thermal polymerisation the resulting materials were crushed, ground and sieved. First generation MIPs were produced in protic polar porogenic solvents (mixture of methanol (MeOH) and acetonitrile (ACN)). These MIPs were used and validated as sorbents for solid phase extraction and binding assays. Second generation MIPs were produced with polar aprotic porogenic solvent (DMSO). These polymers were packed in HPLC columns in order to investigate their molecular recognition properties in a dynamic mode. The study of the mobile phase composition included two major parameters: organic modifier content and pH value. Retention factors illustrate selective binding of the template from the imprinted polymers, compared to structurally related compounds.

Rapid spectrofluorimetric determination of lisinopril in pharmaceutical tablets using sequential injection analysis.

The present work reports for the first time a simple and rapid method for the spectrofluorimetric determination of lisinopril (LSP) in pharmaceutical formulations using sequential injection analysis (SIA). The method is based on reaction of LSP with o-phthalaldehyde (OPA) in the presence of 2-mercaptoethanol (borate buffer medium, pH=10.6). The emission of the derivative is monitored at 455 nm upon excitation at 346 nm. The various chemical and physical conditions that affected the reaction were studied. The calibration curve was linear in the range 0.3-10.0 mg L(-1) LSP, at a sampling rate of 60 injections h(-1). Consumption of OPA reagent was significantly reduced compared with conventional flow injection (FI) systems, because only 50 microL of OPA was consumed per run. The method was found to be adequately precise ( s(r)=2% at 5 mg L(-1) LSP, n=10) and the 3 sigma detection limit was 0.1 mg L(-1). The method was successfully applied to the analysis of two pharmaceutical formulations containing LSP. The results obtained were in good agreement with those obtained by use of high-performance liquid chromatography (HPLC), because the mean relative error, e(r), was <1.8%.

Estimation of the applicability of differential spectroscopic method for the determination of lisinopril in tablets and for the evaluation of its stability.

Classic spectroscopy and first-, second-, third derivative UV-spectroscopy methods were applied to the determination of lisinopril (LIS) in the model solutions, tablets as well as to estimate its stability in solid phase. These results were compared with results received by the HPLC method. Derivative UV-spectroscopy and the HPLC method was tested for: selectivity, precision, linearity, accuracy and repeatability. The study shows that derivative UV-spectroscopy (only first and second derivative) and HPLC can be successfully applied for the quantitative analysis of lisinopril in pharmaceutical formulations and evaluation of its stability in solid phase.