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The serotonin receptor ligands and their related compounds are used for the treatment of central nervous system (CNS)-related disorders. The retention profile of six serotonin receptor ligands such as aripiprazole, ziprasidone, risperidone, olanzapine, mianserin, and quetiapine was investigated on two stationary phases that differ in polarities (C8 alkyl and pentafluorophenylpropyl (PFP)). The design of experiments (DoE) methodology was used to define the retention mechanism and to produce acceptable separation results. The chromatographic conditions that provide the best selectivity characteristics are considered for the separation of structurally related oxidative impurities. The selected conditions (C8 stationary phase, 40% acetonitrile, 20mM ammonium acetate, 25 ℃) showed better sensitivity and improved selectivity in the separation of aripiprazole, ziprasidone, and their oxidative degradants compared to the already developed high-performance liquid chromatography methods. The selected method was validated for the determination of aripiprazole in pharmaceutical forms and was found to be selective, sensitive, precise, accurate, and robust. It can be used as a basis for rapid chromatographic profiling of serotonin receptor ligands and their related impurities.

Background: Since the beginning of the HIV/AIDS epidemic, 75 million people have been infected with the HIV and about 32 million people have died of AIDS. Investigation of the molecular mechanisms critical to the HIV replication cycle led to the identification of potential drug targets for AIDS therapy. One of the most important discoveries is HIV-1 protease, an enzyme that plays an essential role in the replication cycle of HIV. Objective: The aim of the present study is to synthesize and investigate anti-HIV-1 protease activity of some chalcone derivatives with the hope of discovering new lead structure devoid drug resistance. Methods: 20 structurally similar chalcone derivatives were synthesized and their physico-chemical characterization was performed. Binding of chalcones to HIV-1 protease was investigated by fluorimetric assay. Molecular docking studies were conducted to understand the interactions. Results: The obtained results revealed that all compounds showed anti-HIV-1 protease activity. Compound C1 showed the highest inhibitory activity with an IC50 value of 0.001 μM, which is comparable with commercial product Darunavir. Conclusion: It is difficult to provide general principles of inhibitor design. Structural properties of the compounds are not the only consideration; ease of chemical synthesis, low molecular weight, bioavailability, and stability are also of crucial importance. Compared to commercial products the main advantage of compound C1 is the ease of chemical synthesis and low molecular weight. Furthermore, compound C1 has a structure that is different to peptidomimetics, which could contribute to its stability and bioavailability.

The appearance of the face is one of the most important factors influencing our perception of beauty. However, few studies have attempted to quantitate what one perceives as beauty. Therefore, this study was conducted with the goal of providing physicians with anatomical data that demonstrate which facial traits most influence our perception of one's attractiveness. In the first phase of the study, faces of 60 participants (30 males and 30 females) were photographed. Next, the photographs were shown to another group of 120 study members (60 males and 60 females), who evaluated the facial features using a Visual Analogue Scale. The highest rated facial parts were then measured using the ImageJ program. In men, the most attractive parts of the face were Type 1:2 lips, a Type IV nose of medium width, blue eyes, brown hair, and a very narrow face. Among females, the most attractive parts of the face were Type 2:1 lips, Type III and V noses of medium width, dark brown eyes, blonde hair, and a narrow face. This is the first study in which the most aesthetically important facial parts have been comprehensively examined. The results obtained in our study show a higher degree of representativeness compared to other studies due to a different methodological approach and can be used as an aesthetic guide and can help in the planning of aesthetic surgery such as lip augmentation and rhinoplasty. Clin. Anat. 33:214-222, 2020. © 2019 Wiley Periodicals, Inc.

HIV protease inhibitors are one of the most important agents for the treatment of HIV infection. In this work, molecular modeling studies combining 3D-QSAR, molecular docking, MESP, HOMO, and LUMO energy calculations were performed on propiophenone derivatives to explore structure activity relationships and structural requirements for the inhibitory activity. The aim of this study was to create a field point–based 3D-QSAR (3D-Quantitative structure-activity relationship) model by using chalcone structures with anti-HIV-1 protease activity from our previous study and to design new potentially more potent and safer inhibitors. The developed model showed acceptable predictive and descriptive capability as represented by standard statistical parameters R2 (0.94) and Q2 (0.59). High correlation between experimental and predicted activities of training set is noticed. All compounds fit into the defined applicability domain. The derived pharmacophoric features were further supported by MESP and Mulliken charge analysis using density functional theory. Statistically significant variables from 3D-QSAR were used to define key structural characteristics which enhance anti-HIV-1 protease activity. This information has been used to design new structures with anti-HIV-1 protease activity. Docking studies were conducted to understand the interactions in predicted compounds. All the compounds were subjected to in silico ADMET profiling in order to select the best potential drug candidates.

Central composite experimental design was used for fast, simple, and accurate high-performance liquid chromatography (HPLC) determination of hydrochlorothiazide, amlodipine and valsartan in combined dosage forms. This method avoids the disadvantages of the traditional analytical approach, which is time-consuming, involves a large number of runs, and does not allow establishing the multiple interacting parameters. On the basis of preliminary experiments and physicochemical characteristic of analyzed substances, three independent variables (methanol content, pH of the mobile phase, and column temperature) were selected as input, while as dependent variables, six responses (retention time of hydrochlorothiazide, retention time of amlodipine, retention time of valsartan, asymmetry of hydrochlorothiazide peak, asymmetry of amlodipine peak, and asymmetry of valsartan peak) were chosen. Face centered central composite design enables an estimation of factors which have the most importance. After optimizing experimental conditions, a separation was conducted on a Zorbax C-8 (150 mm x 4.6 mm, 5 mu m) column with a mobile phase consisting of methanol-acetonitrile-acetate buffer 40: 20: 40 (v/v/v), pH adjusted to 3.5 with acetic acid, flow rate of 1 mL/min and column temperature of 40 C-deg. The method was successfully applied to the simultaneous separations of these active drug compounds in their commercial dosage forms.

The pKa values of twelve β-hydroxy-β-arylalkanoic acids and ibuprofen were determined using a modified RP-HPLC method. The stationary phase was octadecyl modified (C-18) silica gel, and the mobile phases were mixtures of methanol and one of ten different buffers (60:40 volume ratio). The mean retention time of each compound was plotted against the pH of each of the ten used mobile phases. The inflection point of the obtained sigmoidal curve represents the s w pKa of a compound. Using s w pKa in previously established equations for the specific methanol/buffer mixture, the w w pKa values (in pure water) were calculated. The obtained w w pKa values for the synthesized compounds were in a range from 3.40 to 3.74, and the w w pKa for ibuprofen was 4.27. The Predicted pKa values for this type of compounds in the MarvinSketch 5.11.5. Program were in poor correlation with the experimental results, while in ACD/ /I-Labs pKa values were calculated as a wide range. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. 172041]

Introduction/Objective Biofilm and pyocyanin production are essential components of Pseudomonas aeruginosa virulence and antibiotic resistance. Our objective was to examine inhibitory effect of synthetized propafenone derivatives 3-(2-Fluorophenyl)-1-(2(2-hydroxy-3-propylamino-propoxy)-phenyl)-propan-1-one hydrochloride (5OF) and 3-(2-Trifluoromethyl-phenyl)-1-(2-(2-hydroxy-3-propylamino-propoxy)-phenyl)-propan-1-one hydrochloride (5CF3) on biofilm and pyocyanin in Pseudomonas aeruginosa clinical strains. Methods Effects were tested on nine clinical isolates and one control laboratory strain of P. aeruginosa. In vitro analysis of biofilm growing was performed by incubating bacteria (0.5 McFarland) with 5OF and 5CF3 (500-31.2 µg/ml) and measuring optical density (OD) at 570 nm. Bacteria in medium without compounds were positive control. Blank medium (an uninoculated medium without test compounds) was used as negative control. Pyocyanin production was estimated by OD at 520 nm, after bacteria incubated with 5CF3 and 5OF (250 and 500 mg/ml), treated with chloroform, and chloroform layer mixed with HCl. Results A total of 500 µg/ml of 5OF and 5CF3 completely inhibited biofilm formation in 10/10 and 4/10 strains, respectively. A total of 250 µg/ml of 5OF and 5CF3 strongly inhibited biofilm formation in 7/10 strains, while inhibition with 125 µg/ml of 5OF and 5CF3 was moderate. Lower concentrations had almost no effect on biofilm production. Pyocyanin production was reduced to less than 40% of the control value in 6/9, and less than 50% of the control in 7/9 strains with 500 mg/ml of 5OF and 5CF3, respectively. At 250 µg/ml 5OF and 5CF3, most strains had pyocyanin production above 50% of the control value. Conclusion Synthetized propafenone derivatives, 5OF and 5CF3, inhibited biofilms and pyocyanin production of Pseudomonas aeruginosa clinical strains. Presented results suggest that propafenone derivatives are potential lead-compounds for synthesis of novel antipseudomonal drugs., Uvod/Cilj Proizvodnja biofilma i piocijanina je važan faktor virulencije i antibiotske rezistencije bakterije Pseudomonas aeruginosa. Cilj rada je bio da se ispita inhibitorni efekat sintetisanih propafenonskih derivata, 3-(2-fluoro-fenil)-1-[2-(2-hidroksi-3-propilamino-propoksi)-fenil]-propan-1-on-hidrohlorid) (5OF) i 3-(2-trifluorometilfenil)-1-[2-(2-hidroksi-3-propilamino-propoksi)-fenil]-propan-1-on-hidrohlorid) (5CF3), na produkciju biofilma i piocijanina kod kliničkih izolata bakterije Pseudomonas aeruginosa. Metode Efekat propafenonskih derivata ispitan je na devet kliničkih izolata i jednom standardnom soju bakterije P. aeruginosa. Uticaj na produkciju biofilma ispitan je in vitro, inkubacijom bakterija (0,5 po Makfarlandu) sa 5OF i 5CF3 (500-31,2 µg/ml), i merenjem optičke gustine na 570 nm. Bakterije u medijumu bez ispitivanih jedinjenja su bile pozitivna kontrola, a sam medijum negativna kontrola. Proizvodeni piocijanin, koji je određivan merenjem optičke gustine na 520 nm, na koinkubacije bakterija sa 5CF3 ili 5OF (250 i 500 mg/ml), tretiran je hloroformom i mešanjem hloroformskog sloja sa HCl. Rezultati Pri koncentraciji od 500 µg/ml 5OF je doveo do potpune inhibicije produkcije biofilma kod svih ispitivanih sojeva (10/10). Inhibicija biofilma sa 500 µg/ml 5CF3 bila je potpuna kod 4/10 sojeva. Pri koncentraciji 5OF i 5CF3 od 250 µg/ml produkcija biofilma kod većine ispitanih izolata bila je slaba, dok je pri koncentraciji 125 µg/ml 5OF odnosno 5CF3 produkcija bila umerena. Niže koncentracije 5OF i 5CF3 nisu imale inhibitorni efekat na formiranje biofilma. U prisustvu 500 mg/ml 5OF u 6/10 ispitivanih sojeva produkcija piocijanina pala je na manje od 40% u odnosu na kontrolnu vrednost. Ista koncentracija (500 mg/ml) 5CF3 snizila je produkciju piocijanina na manje od 50% od kontrole u 7/9 sojeva. Pri koncentraciji 250 µg/ ml 5OF ili 5CF3 većina sojeva produkovala je piocijanin iznad 50% u odnosu na pozitivnu kontrolu. Zaključak Sintetisani propafenonski derivati, 5OF i 5CF3, inhibiraju produkciju biofilma i piocijanina kod kliničkih sojeva bakterije Pseudomonas aeruginosa. Dobijeni rezultati ukazuju na to da propafenonski derivati predstavljaju moguća polazna jedinjenja za sintezu novih antipseudomonasnih agenasa.

Retention behaviour of twenty-one chalcones synthesized in our laboratory was tested in three thin-layer chromatography (RP-TLC) systems (acetonitrile–water, ethanol–water and acetone–water) and chromatography parameters $$ {R}_M^0 $$ , S and C0 were calculated. The most suitable RP-TLC system (acetonitrile–water) and chromatography parameter (C0) for lipophilicity prediction of tested compounds were selected on the basis of the highest correlations with calculated logP values. In selected system, compound 12 had the highest, whereas 47 had the lowest C0 value. QSRR analysis was performed and three models representing relationships between C0 and selected molecular descriptors were created—MLR(C0), PLS(C0) and SVM(C0). Interpretation of molecular descriptors which form statistically the most reliable SVM(C0) model identified the most important structural and physico-chemical properties that influence retention behaviour of tested compounds. In addition, descriptors with the highest influence on $$ {R}_M^0 $$ as well as on C0 calculated in the remaining two RP-TLC systems were identified and interpreted.

One of the most commonly used surfactants in the production of split virus influenza vaccine is nonionic surfactant Triton X-100. After splitting of the virus is accomplished, Triton X-100 is removed from the vaccine by subsequent production steps. Because of toxicity of Triton X-100, which remains in the vaccine in residual amounts, a sufficiently sensitive method for its detection and quantification needs to be defined. Two methods for determination of Triton X-100 residuals were developed: the UV-spectrophotometry and HPLC methods. For both methods, preparation of vaccine samples and removal of proteins and virus particles were crucial: samples were treated with methanol (1:1) and then centrifuged at 25 000 × g for 30 min. After such treatment, the majority of vaccine components that interfered in the UV region were removed, and diluted samples could be directly measured. The chromatographic system included C18 column, step methanol gradient, and detection at 225 nm with a single peak of Triton X-100 at 12.6 min. Both methods were validated and gave satisfactory results for accuracy, precision, specificity, linearity, and robustness. LOQ was slightly lower for the HPLC method. Hence, it was shown that both methods are suitable for analysis of residual amounts of Triton X-100, with the advantages of the UV method being its simplicity and availability in most laboratories.

An imbalance between oxidants and antioxidants in favour of oxidants, potentially leading to damage, is termed oxidative stress. Antioxidants (AO), either enzymatic or non-enzymatic, are the ones that can reduce diverse effects of pro-oxidants such as DNA, proteins and lipids damage. Chalcones (1,3-diaryl-2-propen- 1-ones) are open chain flavonoids that are widely biosynthesized in plants. Aim of this study was to test antioxidative potency of 15 chalcones (Chs) in in vitro model in serum (native conditions), so as with exogenously induced oxidative stress. Material and methods: Oxidative stress was induced in serum samples of healthy individuals with 0.25 mmol/L terc-buthyl-hydroperoxide (TBH), and then we monitored the effects of various concentrations of chalcones on oxidative stress parameters: total antioxidative status (TAS), total oxidative status (TOS), total concentration of sulfhydryl group (SHG) and prooxidative-antioxidative balance (PAB), and calculated prooxidative score, antioxidative score, and oxy score (OS). Nonparametric repeated measures ANOVA (Friedman's test) was used for comparison of antioxidative potency of samples with 15 different chalcones, in a native state and upon TBH influence. Spearman's nonparametric correlation analysis was used for estimation of relation between different parameters. Results: Negative Oxy Score (OS) values for Chs11-15 showed significantly stronger antioxidative potency compared to other investigated chalcones (p < 0.05). Ch11, Ch13 and Ch14 remained with negative OS even after TBH addition, whereas OS of Ch12 and Ch15 became positive, with small nominal values. Samples with Ch11 and Ch13 showed significant negative correlation between TAS and TOS (p < 0.05 for both), but in Ch14 sample the negative correlation existed between TAS and PAB (p < 0.05). Conclusion: Lower value of OS (i.e. better antioxidative potency) was noticed in samples with Ch11-Ch15. Electron-donor effects of substituent groups as a structural part of these chalcones could explain its antioxidative capability. Phenolic and methyl groups are responsible for antioxidative ability enhancement of five chalcones with the best activity.

Backgroung/Aim. Since combining conventional drugs with herbal medicinal products is in current research focus and possible of great interest as therapy improvement way, the aim of this study was to determine the effects of well-established antiatherosclerotic drug atorvastatin (CAS number 134523-00-5) and commercially available artichoke leaf tincture (ALTINC), used as combined therapy, as well as to compare effects of these two treatments separately. Methods. Experimental animals were divided into five groups: the group I (the control group of rats fed with standard diet during 11 weeks), and the remaining 4 groups of rats (II, III, IV and V) fed with standard diet during the first week and then with hypercholesterolemic diet during the next 10 weeks. The group II of rats were left without treatment, while in the groups III, IV and V were rats treated per os with atorvastatin (1.15 mg/kg body wright - b.w.), ALTINC (0.1 mL/kg b.w.) and their combination in same doses, respectively, for the last six weeks. Results. The cholesterol rich diet led to pronounced hyperlipidemia which could not be overcame with the therapy. However, the therapy showed positive effects on abdominal aorta wall thickness and parameters of oxidative stress (malondialdehyde - MDA, proxidative-antioxidative balance - PAB) and anti- oxidative protection (reduced glutathione - GSH, paraoxanase 1 - PON1, superoxide dismutase - SODA SH groups), especially ALTINC was successful in oxidative status improvement. Conclusion. Separate treatments comparison showed that artichoke leaf tincture is very potent antioxidant with beneficial effects in early stages of atherosclerosis. Since atorvastatin and constituents of ALTINC probably have different mechanisms of action, simultaneous use of both therapies could be beneficial but should be further investigated since our results showed that ALTINC is less effective when used in combination with atorvastatin. Uvod/Cilj. Savremena istraživanja sve više se okreću mogućnosti kombinovanja konvencionalne terapije sa biljnim lekovitim proizvodima. Cilj ovog istraživanja bio je praćenje efekata atorvastatina (CAS broj 134523-00-5), poznatog leka koji se koristi u terapiji hiperlipidemije, i tinkture lista artičoke primenjenih pojedinačno, kao i u obliku kombinovane terapije. Metode. Eksperimentalne životinje bile su podeljene u pet grupa: grupa I bila je kontrolna i činili su je pacovi hranjeni standardnom hranom za glodare tokom 11 nedelja, dok su preostale četiri grupe (II-V) činili pacovi koji su hranjeni standardnom hranom tokom prve nedelje, a potom u narednih 10 hranom bogatom holesterolom. Grupa II nije dobijala nikakav tretman, dok su III, IV i V posle 4. nedelje od početka uzimanja hrane bogate holesterolom, tokom narednih šest nedelja tretirane per os, redom: atorvastatinom u dozi od 1,15 mg/kg, tinkturom artičoke u dozi od 0,1 mL/kg i njihovom kombinacijom u istim dozama. Rezultati. Hrana bogata holesterolom izazvala je uznapredovalu hiperholesterolemiju na koju terapija nije značajno delovala. Terapija je pokazala povoljan efekat na debljinu zida trbušne aorte, parametre oksidativnog stresa (malondialdehid - MDA, prooksidativni-antioksidativni balans - PAB) i anti-oksidativne zaštite (redukovani glutation - GSH, SH grupe, paraoksonazu1 - PON1, superoksid dismutazu - SOD). Tinktura lista artičoke pokazala se najboljom u poboljšanju oksidativnog statusa. Zaključak. Poređenje pojedinačne terapije pokazalo je da tinktura lista artičoke ima jako anti-oksidativno dejstvo i pokazuje povoljne efekte u početnim fazama aterosklerotskog procesa. S obzirom na to da atorvastatin i komponente tinkture lista artičoke verovatno deluju različitim mehanizmima, kombinovana primena može imati povoljne efekte, ali se mora dodatno ispitati jer dobijeni rezultati ukazuju da je tinktura lista artičoke manje efikasan kada se primenjuje zajedno sa atorvastatinom.

Context: Polyphenols and flavonoids in artichoke leaf tincture (ALT) protect cells against oxidative damage. Objectives: We examined ALT effects on deoxyribonucleic acid (DNA) damage and lipid profiles in rat plasma and gene expression in rat aorta [haemeoxygenase-1 (HO1), haemeoxygenase-2 (HO2), NADPH oxidase 4 (NOX-4), monocyte chemoattractant protein-1 (MCP-1) and nuclear factor (erythroid-derived 2)-like 2 (Nrf2)]. Materials and methods: Eighteen male Wistar albino rats were divided into three groups (n = 6/group): The control group (CG) was fed with standard pellet chow for 11 weeks; the AD group was fed for a similar period of time with pellet chow supplemented with 2% cholesterol, 3% sunflower oil and 1% sodium cholate. The ADA group was fed with pellet chow (for 1 week), the atherogenic diet (see above) for the following 4 weeks and then with ALT (0.1 mL/kg body weight) and atherogenic diet for 6 weeks. According to HPLC analysis, the isolated main compounds in ALT were chlorogenic acid, caffeic acid, isoquercitrin and rutin. Results: Normalized HO-1 [0.11 (0.04–0.24)] and MCP-1 [0.29 (0.21–0.47)] mRNA levels and DNA scores [12.50 (4.50–36.50)] were significantly lower in the ADA group than in the AD group [0.84 (0.35–2.51)], p = 0.021 for HO-1 [0.85 (0.61–3.45)], p = 0.047 for MCP-1 and [176.5 (66.50–221.25)], p = 0.020 for DNA scores. HO-1 mRNA was lower in the ADA group than in the CG group [0.30 (0.21–0.71), p = 0.049]. Conclusions: Supplementation with ALT limited the effects of the atherogenic diet through reduced MCP-1 expression, thereby preventing oxidative damage.

To protect organisms from ionizing radiation (IR), and to reduce morbidity or mortality, various agents, called radioprotectors, have been utilized. Because radiation-induced cellular damage is attributed primarily to the harmful effects of free radicals, molecules with radical-scavenging properties are particularly promising as radioprotectors. Early development of such agents focused on thiol synthetic compounds, known as WR protectors, but only amifostine (WR-2721) has been used in clinical trials as an officially approved radioprotector. Besides thiol compounds, various compounds with different chemical structure were investigated, but an ideal radioprotector has not been found yet. Plants and natural products have been evaluated as promising sources of radioprotectors because of their low toxicity, although they exhibit an inferior protection level compared to synthetic thiol compounds. Active plant constituents seem to exert the radioprotection through antioxidant and free radical-scavenging activities. Our research established that plants containing polyphenolic compounds (raspberry, blueberry, strawberry, grape, etc.) exhibit antioxidative activities and protect genetic material from IR.

The aim of the present study was to optimize a chromatographic method for the analysis of atorvastatin (acid and lactone forms), ortho- and para-hydroxyatorvastatin by using an experimental design approach. Optimization experiments were conducted through a process of screening and optimization. The purpose of a screening design is to identify the factors that have significant effects on the selected chromatographic responses, and for this purpose a full 23 factorial design was used. The location of the true optimum was established by applying Derringer's desirability function, which provides simultaneously optimization of all seven responses. The ranges of the independent variables used for the optimization were content of acetonitrile in mobile phase (60-70%), temperature of column (30-40 C degrees) and flow rate (0.8-1.2 mL min(-1)). The influences of these independent variables were evaluated for the output responses: retention time of first peak (p-hydroxyatorvastatin) and of last peak (atorvastatin, lactone form), symmetries of all four peaks and relative retention time of p-hydroxyatorvastatin. The primary goal of this investigation was establishing a new simple and sensitive method that could be used in analysis of biological samples. The method was validated and successfully applied for determination of atorvastatin (acid and lactone forms) and its metabolites in plasma.

Gastrointestinal absorption of thirteen novel beta-hydroxy-beta-arylalkanoic acids (HAA) with anti-inflammatory activity was predicted by use of biopartitioning micellar chromatography and compared to ibuprofen. All tested HAA have lower retention factors (k) and lower expected gastrointestinal absorption than ibuprofen, whereas derivatives with the highest values of k are 1C, 2APTF and 2C. Quantitative structure-retention relationship (QSRR) analysis was performed in order to identify molecular descriptors with the highest influence on k and ANN(k) model was selected as optimal. Descriptors which form this model (nBM, P VSA_LogP_8 and Eta_L) indicate that replacement of phenyl ring with a saturated or partially unsaturated one, as well as presence of halogens and nitro group should positively affect k values. On the basis of these conclusions, six novel HAA were designed and selected QSRR model was used for the prediction of their k values.

An experimental design method was used for fast, simple, and accurate high-performance-pressure liquid chromatograpy (HPLC) determination of losartan and hydrochlorothiazide in combined dosage forms. This method avoids the disadvantages of the traditional analytical approach, which is time-consuming, involves a large number of runs, and does not allow the determination of multiple interacting parameters. On the basis of preliminary experiments, three independent variables (methanol content, pH value of the mobile phase, and flow rate) were selected as input, and as dependent variables, five responses (retention time of hydrochlorothiazide, retention time of losartan, asymmetry of hydrochlorothiazide peak, asymmetry of losartan peak, and resolution) were chosen. A full 2(3) factorial design was used to determine which factors had an effect on the studied response. Afterwards, experimental design was used to optimize these influencing parameters in the previously selected experimental domain. After optimizing the experimental conditions, a separation was conducted on a Zorbax C-8 (150 mm x 4.6 mm, 5 mu m particle size) column with a mobile phase consisting of methanol-acetonitrile-acetate buffer 45:20:35 v/v/v, pH 4.8 with flow rate of 0.82 mL min(-1) and column temperature of 25 A degrees C. The developed method was successfully applied to simultaneous separation of these active drug compounds in their commercial pharmaceutical dosage forms.

Two flavonoids, rutin and hesperidin, were investigated in vitro for anticoagulant activity through coagulation tests: activated partial thromboplastin time (aPTT), prothrombin time (PT) and thrombin time (TT). Only an ethanolic solution of rutin at the concentration of 830 mu M prolonged aPTT, while TT and PT were unaffected. In order to evaluate whether the prolongation of aPTT was due to the decrease of coagulation factors, the experiment with deficient plasma was performed, showing the effects on factors VIII and IX. Since pharmacological activity of flavonoids is believed to increase when they are coordinated with metal ions, complexes of these flavonoids with Al(III) and Cu(II) ions were also tested. The results showed that complexes significantly prolonged aPTT and had no effects on PT and TT. Assay with deficient plasma (plasma having the investigated factor at less then 1%) revealed that complexes could bind to the coagulation factors, what may lead to a non-specific inhibition and aPTT prolongation. An effort was made to correlate stability of complexes with their anticoagulant properties.

In this work, the behavior of the active pharmaceutical substances methylprednisolone (in a form of methylprednisolone sodium succinate) in finished pharmaceutical dosage form, i.e., freeze-dried powder for injections, was examined. The goal was to evaluate the chemical stabilities of methylprednisolone sodium succinate packaged in a dual chamber vial, as a specific container closure system. The effect of different parameters: temperature, moisture and light were monitored. The method proposed by United States Pharmacopeia was used to determine concentrations of methylprednisolone, as the sum of the concentration of methylprednisolone esters (17-hydrogen succinate and 21-hydrogen succinate) and free methylprednisolone. The HPLC method was used for stability evaluation of the active substance and determination of related substances. Four main degradation products were registered. Temperature has a major impact on the degradation process with the appearance of 3 degradation products (impurities B, C and D), while the presence of light caused an increasing content of impurity A. Identification of impurity B, C and D has been realized using mass and NMR spectroscopy. All three substances are substances related to methylprednisolone. U ovom radu ispitivane su osobine farmakološki aktivne supstance metilprednizolona (u obliku metilprednizolon-natrijum-sukcinata) u gotovom proizvodu - liofilizatu za rastvor za injekcije. Cilj rada je ispitivanje hemijske stabilnosti metilpredni-zolon-natrijum-sukcinata u dvokomornoj bočici, kao specifičnom sistemu kontaktnog pakovanja. Ispitan je efekat različitih parametara: temperature, vlage i svetlosti. Za određivanje koncentracije metilprednizolona, kao zbirne koncentracije metilprednizolon estara (17-hidrogen-sukcinata i 21-hidrogen-sukcinata) i slobodnog metilprednizolona, korišćena je metoda opisana u Američkoj farmakopeji. Za ispitivanje srodnih supstanci primenjena je HPLC metoda. Uočena su 4 degradaciona proizvoda. Dokazano je da povećanje temperature ima najveći značaj na proces degradacije i utiče na povećanje sadržaja nečistoća B, C i D, dok prisustvo svetlosti dovodi do povećanja sadržaja nečistoće A. Nečistoće B, C i D su identifikovane primenom masene i NMR spektroskopije. Sve tri nečistoće su identifikovane kao srodne supstance metilprednizolona.

A 3D-QSAR model using the GRIND/ALMOND descriptors has been performed on a set of 49 angiotensin receptor blockers, also known as angiotensin II receptor antagonists, a family of agents that bind to and inhibit the angiotensin II type 1 receptor. The most commonly used chemical probes: DRY (hydrophobic interaction), O (carbonyl oxygen sp 2, hydrogen bond donor), N1 (NH neutral, hydrogen bond acceptor) and TIP (shape descriptor molecular forms) were derived from the GRID molecular interaction fields. A statistical approach was undertaken using the method of partial least squares within the Pentacle program. The results show satisfactory accuracy of the prediction model (RMSEE = 0.239, R 2 = 0.94, Q 2 = 0.85). The V597 (DRY-TIP) and V763 (O-TIP) represent the most significant variables that correlate positively with the activity of the ARBs. Thirty novel structures of ARBs were designed, according to the developed 3D-QSAR model and pharmacophore, what might set the basis for development of new antihypertensive drugs.

The aim of this study was to investigate the encapsulation of α -lipoic acid (LA) onto chitosan microbeads in order to achieve a sustained release. Microbeads of different compositions were synthetized by inverse emulsion method, and characterized by in vitro swelling and drug release studies, FT-IR, DSC and SEM analyses. The encapsulation of LA was performed by swelling of previously synthetized dry microbeads in LA solution. The encapsulation efficiency was determined in terms of LA concentration, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The swelling of synthetized microbeads was well pronounced in the acidic media. FT-IR analysis confirmed the presence of LA in microbeads and its interactions with matrices, which was in agreement with the results revealed by DSC analysis. SEM analysis revealed the porous structure, micron-size and spherical shape of the synthetized beads. Encapsulation efficiency was in the range from 46.8 to 58.5% depending on the hydrogel composition. The microbeads exhibited sustained release of LA under conditions mimicking gastrointestinal tract. Results of antioxidant activity of released LA showed good correlation with the concentration of the released LA. The obtained results suggested that synthetized microbeads have promising potential for oral administration with reduced dosing frequency in comparison to the immediate release formulations.

A simple and sensitive liquid chromatography-tandem mass spectrometry method was developed for the quantification of atorvastatin, ortho-hydroxyatorvastatin, para-hydroxyatorvastatin, and atorvastatin lactone in rat plasma. Solid-phase extraction was used for preparation of samples. Rosuvastatin was chosen as an internal standard. Chromatographic separation was achieved on ZORBAX Eclipse C-18 Analytical, 4.6 x 100 mm (3.5 mu m) column with a gradient mobile phase composed of acetonitrile and 0.1% acetic acid, at a flow rate of 400 mu L min(-1). The column was kept at constant temperature (25 degrees C), and autosampler tray temperature was set at 4 degrees C. The following selected reaction monitoring (SRM) transitions were selected: (m/z, Q1 - gt Q3, collision energy) atorvastatin (559.47 - gt 440.03, 22 eV), atorvastatin lactone (541.36 - gt 448.02, 19 eV), orthohydroxyatorvastatin (575.20 - gt 440.18, 20 eV), para-hydroxyatorvastatin (575.54 - gt 440.18, 20 eV), and rosuvastatin (482.25 with selected combination of two fragments 257.77, 31 eV, and 299.81, 35 eV) in positive ion mode. The method was validated over a concentration range of 0.5-20 ng mL(-1) for ortho-hydroxyatorvastatin and para-hydroxyatorvastatin and 0.1-20 ng mL(-1) for atorvastatin and atorvastatin lactone with excellent linearity (r(2) gt = 0.99). This method demonstrated acceptable precision and accuracy at four quality control concentration levels. The detection limits were 0.1 and 0.13 ng mL(-1) for orthohydroxyatorvastatin and para-hydroxyatorvastatin, respectively, and 0.05 ng mL(-1) for atorvastatin and atorvastatin lactone. All analytes were found to be stable at examined conditions. Validated method was applied for determination of atorvastatin and its metabolites in plasma of experimental animals.

A rapid and sensitive assay for quantitative determination of rutin in oral dosage forms based on isocratic reversed phase high performance liquid chromatography (RP-HPLC) was developed and validated. Using a C-18 reverse-phase analytical column, the following conditions were chosen as optimal: mobile phase methanol-water 1:1 (v/v), pH 2.8 (adjusted with phosphoric acid), flow rate = 1 mL min(-1) and temperature T = 40.0 degrees C. Linearity was observed in the concentration range 8-120 mu g mL(-1) with a correlation coefficient of 0.99982 and the limit of detection (LOD) = 2.6 mu g mL(-1), and limit of quantification (LOQ) = 8.0 mu g mL(-1). Intra- and inter-day precision were within acceptable limits. Robustness test indicated that the mobile phase composition and pH influence mainly the separation. The proposed method allowed direct determination of rutin in pharmaceutical dosage forms in the presence of excipients, but is not suitable for preparations where compounds structurally/chemically related to rutin may be present.

The current research provides a simplified sample preparation procedure for the accurate estimation of quercetin in pure and in the pharmaceutical dosage form. Direct spectrophotometric method for the determination of quercetin in the presence of ascorbic acid was established. The influences of medium, wavelength, pH, temperature and the ionic strengths on quercetin determination were investigated. The best conditions for calibration curve are: 50% ethanol, λ = 370 nm, pH = 4.2, T = 34 °C and I = 7.5 × 10 –5 M. Beer’s law is obeyed in the concentration range 1.0–12.0 μg ml –1 for quercetin. The corresponding detection limit is 0.76 μg ml –1 . The proposed method was verified by analyzing Quercetin + C capsules, Twinlab ® .

In this paper, influence of temperature and light on stability of montelukast in solutions (standard substance, chewable tablets and film coated tablets) which were prepared during routine analysis of related substances was investigated. The content of montelukast and degradation products was determined by HPLC method at defined time intervals (influence of temperature was tested at the beginning, after 24 hours, and after 48 hours, and influence of light was tested at the beginning of the experiment and after 12 hours). Stability of montelukast in tested solutions under influence of temperature was acceptable. The content of montelukast ranged from 98 % to100 %, which meets the pharmacopoeia requirements and specifications. Individual and total impurities were within acceptable limits. Presence of photosensitive functional groups in structure of montelukast led to significant photodegradation. Individual and total impurities in samples which were under influence of light were above acceptable limits. Photodegradation occured during first 12 hours. Content of MOK-3 sulphoxide and unknown impurity on RRT 0.76 was above limits in all examined samples. Over a period of 12 h the content of those impurities grew 8-40 fold compared to time zero values. U radu je dat prikaz ispitivanja uticaja temperature i svetlosti na stabilnost montelukasta u rastvorima (standardne supstance, tabletama za žvakanje i filmom obloženih tableta) koji se pripremaju u rutinskoj analizi u toku postupka ispitivanja prisustva srodnih supstanci. Sadržaj montelukasta i nastalih degradacionih proizvoda praćen je primenom RP-HPLC metode u definisanim vremenskim intervalima (0, 24 i 48h kada je u pitanju temperatura, odnosno 0 i 12h kada je u pitanju svetlost). Rezultati ispitivanja uticaja temperature, kao spoljašnjeg faktora nestabilnosti, ukazuju da su analizirani rastvori standarda montelukasta, tableta za žvakanje i filmom obloženih tableta stabilni u prihvatljivim granicama. Sadržaj montelukasta se kreće od 98 % - 100 % što zadovoljava farmakopejske zahteve, kao i zahteve specifikacije proizvoda. Pojedinačne i ukupne nečistoće su u dozvoljenim granicama. Prisustvo fotoreaktivnih funkcionalnih grupa u strukturi montelukasta uslovilo je nastanak fotodegradacionih proizvoda u količinama koje su izvan granica definisanih u monografiji montelukasta, kao i granica definisanih u specifikaciji gotovog proizvoda. Do fotodegradacije dolazi već u toku prvih 12 h. Porast sadržaja MOK-3 sulfoksida i nečistoće sa relativnim retencionim vremenom oko 0,76 zabeležen je u sva tri ispitivana uzorka. U vremenskom intervalu od 12 h uočen je porast sadržaja navedenih nečistoća 8-40 puta u poređenju sa vrednostima u početnom vremenu ispitivanja.

This paper describes a simple densitometric method for the determination of metroprolol tartrate in tablets and ampoules. After separation on silica gel GF254 plates, using acetone-methanol-triethylamine as the mobile phase for the tablets and acetone-triethylamine for ampoules, the chromatographic zones corresponding to the spots of metoprolol were scanned. Quantitation was performed using a computer-controlled Camag TLC scanner and applying five-point calibration with polynomial regression. The calibration function was established in the ranges 1-28 micrograms for tablets and 1-9 micrograms for ampoules. The results obtained are precise and reproducible, with recovery values of 99.1-99.4%.

A method was developed to determine lindane (gamma-HCH), the widely used organochlorine pesticide, and its impurity alpha-HCH in bulk and pharmaceutical products (shampoo, gel and emulsion). The system was able to successfully resolve the lindane peak from alpha-HCH and interfering components. The method displays an excellent linearity over the concentration range 0.5-15.0 mu g mL(-1) for lindane and 0.005-0.15 mu g mL(-1) for alpha-HCH and a precision better than 2.5% from intra- and inter-day analyses. LOQ and LOD were determined to be 0.45 and 0.15 mu g mL(-1) for lindane and 0.005 and 0.002 mu g mL(-1) for alpha-HCH.

A rapid and simple liquid chromatographic method with UV detection has been developed for the determination of hydrochlorothiazide (HCTZ), cilazapril (CL) and its active metabolite cilazaprilat (CLT) in urine. Sample preparation for urine consisted of solid-phase extraction using styrene-divinylbenzene (SDB-2) cartridges. The chromatographic system was a Zorbax Eclipse XDB-C-18 column with a mixture of methanol and 10 mM phosphate buffer, pH 2.3 with gradient (20 to 60% of methanol) as mobile phase at a flow rate of 1.0 mL min(-1). The detection was performed at the wavelength of 206 nm. Enalapril maleat was used as an internal standard. The detector response was linear in the range of 2.4-30.0, 1.6-15.0 and 1.8-20.0 mu g mL(-1) for HCTZ, CL and CLT, respectively. LOQ was determined to be 2.4, 1.6 and 1.8 mu g mL(-1) for HCTZ, CL and CLT, respectively. Both intra- and inter-day precision were within acceptable limits. The method has been applied to urine samples obtained from three hypertensive patients after intake of HCTZ and CL therapeutic dose.

A method for separation of three antidepressants, maprotiline, desipramine, and moclobemide, by reversed-phase high-performance liquid chromatography (RP-HPLC) was developed and validated. To find optimal conditions and estimate the impact of individual parameters on the separation, a complete set of 23 interdependent relationships of the mobile phase composition, temperature, and the volume flow rate were examined. Full separation of the investigated components from a laboratory mixture was achieved on a Supelcosil LC-18 (120 mm × 4.6 mm, 5 µm) column, using two solvent systems, 3% ammonium ion in water/ethanol and acetonitrile, and alternating isocratic gradient–isocratic elution modes. Relevance of the proposed method for therapeutic drug monitoring is anticipated.

A method of Capillary Zone Electrophoresis (CZE) for separation of three most-frequently prescribed antidepressants in the market: maprotiline, desipramine, and moclobemide was developed. The proposed method is fully validated for a ternary laboratory mixture but it is also suitable for individual determination of the investigated components in pharmaceutical dosage forms. Since the preliminary investigations did not show complete separation because of close migration time of desipramine and maprotiline, a complete set, 2(3) experimental design was applied. All the factors that affect separation as well as their mutual interactions were investigated. Voltage, temperature of capillary and the pH of phosphate buffer were independent variables or factors to be investigated in two levels. Applying response surface methodology, from experimental points the appropriate graphs were constructed and optimal chromatographic conditions for the separation were defined. The optimum conditions were: running voltage of 20kV, phosphate buffer (pH = 2.35), temperature 25 degrees C, and UV detection at 200nm. An uncoated (fused silica) capillary (50 mu m i.d.) with a total length of 50cm and a distance of 47cm between the injection and detection points was used.

A method for chromatographic separation of the enantiomers of (±)-metoprolol tartrate (±MeT) employing a chiral mobile phase additive (CMPA) is described. By using silica gel plates previously impregnated with the mobile phase (ethanol-water, 70 + 30, v/v ) containing D -(−)-tartaric acid as chiral selector, direct separation of the enantiomers of ±MeT was achieved. The results of experiments with different concentrations of D -(−)-tartaric acid (5.8, 11.6, and 23 mmol L −1 ) revealed that the best resolution of the enantiomers of ±MeT was achieved with 11.6 mm D -(−)-tartaric acid in both the mobile phase and the impregnation solution, at 25 ± 2°C.Spot visualization on chromatograms was performed by use of a fixed-wavelength ( λ = 254 nm) ultraviolet lamp or an iodine vapor-saturated chamber.

An HPLC method was developed for the determination of S-carboxymethyl-L-cysteine, methylparaben and related compounds in cough syrup preparations. The HPLC operating conditions used were acetonitrile-methanol-water (120:280:600, V/V/V) pH adjusted to 4.0 with 0.1 M HCl, on a 5um Spherisorb ODS2 column (250 mm x 4.6 mm) with a flow rate of 1 mL min−1. The injection volume was 10 μL and absorbance was monitored at 210 nm with 0.05 a.u.f.s. sensitivity.

The inclusion complex formation between Metoprolol tartarata (MeT) and beta -cyclodextrin (beta -CD) has been investigated using hyperchromic shift at lambda (max) 274.4 nm of MeT. Different parameters such as stirring time, solvent composition (aqueous and aqueous/methanol solutions with methanol content up to 50%), pH values 4.0 and 8.0 were established for optimal inclusion complex formation and confirmed two stoichiometric compositions 1:1 and 1:2. Preliminary data. on usage of MeT/beta -CD complex in reversed-phase HPLC indicate the potential application of this complex as a kind of pre-column derivatization for enantiomeric separation of beta (1)-blockers.

Fluocortolone and its esters are synthetic corticosteroids used topically in the treatment of various skin disorders. A method that can be successfully used for the separation and determination of fluocortolone pivalate and fluocortolone hexanoate in suppositories was developed. This method is based on reverse-phase HPLC on Supelcosil LC-18 (25 cm x 4.6 mm, 5 microns), using methanol-acetonitrile-water-glacial acetic acid (17:46:37:0.4 v/v/v/v) as mobile phase at a flow rate of 3.0 ml/min. Detection was carried out using a UV detector at 238 nm. The method developed was validated, and calibration curves were established dependent on peak area. The validated ranges for fluocortolone pivalate and fluocortolone hexanoate are 15-305 micrograms/ml (r = 0.9995) and 15-315 micrograms/ml (r = 0.9996), respectively. The limits of detection and the limits of quantification for both esters were also determined.

A sensitive, accurate and reproducible high performance liquid chromatographic (HPLC) procedure has been developed for the separation and analysis of lorazepam and related compounds. A 20-muL sample was separated by reversed-phase HPLC on a 150 mm x 4.6 mm, 5-mum particle, Wakosil column with 0.1 M ammonium acetate (pH adjusted to 6.0 with acetic acid)-acetonitrile-methanol, 1:1:1 (v/v) as mobile phase.