High-pressure Liquid Chromatographic Method Research Articles

Development and Validation of a Novel RP-HPLC Method for the Analysis of Reduced Glutathione

The objective of this study was the development, optimization, and validation of a novel reverse-phase high-pressure liquid chromatography (RP-HPLC) method for the quantification of reduced glutathione in pharmaceutical formulations utilizing simple UV detection. The separation utilized a C18 column at room temperature and UV absorption was measured at 215 nm. The mobile phase was an isocratic flow of a 50/50 (v/v) mixture of water (pH 7.0) and acetonitrile flowing at 1.0 mL/min. Validation of the method assessed the methods ability in seven categories: linearity, range, limit of detection, limit of quantification, accuracy, precision, and selectivity. Analysis of the system suitability showed acceptable levels of suitability in all categories. Likewise, the method displayed an acceptable degree of linearity (r(2) = 0.9994) over a concentration range of 2.5-60 µg/mL. The detection limit and quantification limit were 0.6 and 1.8 µg/mL respectively. The percent recovery of the method was 98.80-100.79%. Following validation the method was employed in the determination of glutathione in pharmaceutical formulations in the form of a conjugate and a nanoparticle. The proposed method offers a simple, accurate, and inexpensive way to quantify reduced glutathione.

Effect of chemical and microbial vitamin B12 analogues on production of vitamin B12

Strain improvement by genetic manipulation or optimization of fermentation conditions for overproduction of vitamin B(12) has a drawback due to feed back inhibition. To resist the feed back inhibition by analogues of vitamin B(12) in Propionibacterium freudenrechii subsps. shermanii (OLP-5), we have tested with microbially separated B(12) analogues from three different strains. Microbial analogues were differentiated from commercially available vitamin B(12) by high pressure liquid chromatography and spectrophotometric method. An analogue isolated from NRRL-B-4327 was shown to increase vitamin B(12) concentration from 18.53 ± 0.15 to 31.67 ± 0.58 mg/l in OLP-5 strain. The presence of chemical analogue (ICH(2) Co(DH)(2) (H(2)Py)(4)) increased vitamin B(12) production from 16.13 ± 0.15 to 18.53 ± 0.15 mg/l in OLP-5. These findings revealed that addition of B(12) analogues in fermentation media have developed strain resistance to feed back inhibition by vitamin B(12).

Rapid Isocratic HPLC Method and Sample Extraction Procedures for Measuring Carotenoid, Retinoid, and Tocopherol Concentrations in Human Blood and Breast Milk for Intervention Studies

Vitamin A deficiency continues to be a major public health concern in the developing world effecting over 200 million people. Interventions including carotenoid-rich fruits and vegetables, containing α- and β-carotene, and β-cryptoxanthin, are being tested for their potential to alleviate vitamin A deficiency in several countries. Thus, there is a need for a faster isocratic reverse-phase high pressure liquid chromatography (HPLC) method than the methods currently available. Efficient extraction procedures for biological samples are also valuable to save time, materials, and cost. We developed an HPLC method and compared several extraction methods for carotenoids, retinoids, and vitamin E in plasma and human breast milk. The method uses an Agilent 1100 system equipped with a Waters Spherisorb ODS2 column (3 × 125 mm 3 μm) and similar guard column (ThermoScientific ODS2 3 × 20 mm 3 μm). Mobile phase of 7:2:1 acetonitrile:dichloromethane:methanol was run at 0.5 mL min−1, and run times were complete in approximately 8 min. This rapid method uses little solvent and provides excellent results in the analysis of these fat-soluble nutrients. Analyte measurements were repeatable using standards and plasma and were accurate compared to NIST certified and reference serum values. The validated HPLC method was applied to the analysis of plasma samples from a vitamin A intervention study.

Method development and validation for the simultaneous determination of fexofenadine hydrochloride and montelukast sodium using rp-hplc

–This research paper describes simple, specific, accurate and precise reverse phase high pressure liquid chromatographic method for the simultaneous determination of fexofenadine hydrochloride and Montelukast sodium in combined dosage form. The separation was carried out on agilent T.C – C18 column(2)4.6×250m.m with 5 μm internal diameter using acetonitrile: TEA (P H 6) (80:20v/v) as mobile phase at the flow rate of 1ml/min. RP-HPLC separation of the two drugs was carried out in the absorbance mode at 220 nm. The drugs were resolved satisfactorily with Rt values of 3.21 ± 0.01 and 6.66 ± 0.01 for FEX and MTKT, respectively. The linear regression analysis data for the calibration plots showed good linear relationship with r 2 =0.9996 and 0.9998 for FEX and MTKT, respectively in the concentration range of 12144 μg/ml for FEX and 1-12μg/ml for MTKT. The method was validated for precision, robustness, specificity and accuracy. The limit of detection and quantitation were 1.41 and 4.29 μg/ml, respectively for FEX and 0.02 and 0.06 μg/ml, respectively for MTKT. The proposed developed RP-HPLC method can be applied for identification and quantitative determination of FEX and MTKT in bulk drug and drug formulation. Keywords––fexofenadine hydrochloride; montelukast sodium; RP-HPLC; validation.

Diagnostical significance of dimethylarginine in the development of hepatorenal syndrome in patients with alcoholic liver cirrhosis

Chronic consumption of alcohol during a longer period of time leads to the development of cirrhosis with the reduction in metabolic liver function and disorders in arginine metabolism. Hepatorenal syndrome (HRS) is the most severe complication of alcoholic liver cirrhosis. The aim of the study was to analyze disorders in arginine metabolism by monitoring concentrations of asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) in patients with liver cirrhosis and HRS. The study included three groups of subjects: a group of patients with cirrhosis and HRS (24 patients), a group of patients with cirrhosis without HRS (18 patients) and a control group composed of 42 healthy voluntary blood donors. Concentrations of ADMA, SDMA and L-arginine in plasma were measured in all groups using the high pressure liquid chromatography (HPLC) method. The concentration of SDMA was significantly higher in the patients with HRS compared to the patients without HRS and it was also higher than the values obtained from the healthy participants 1.76 +/- 0.3 micromol/L; 1.01 +/- 0.32 and 0.520 +/- 0.18 micromol/L, respectively; p < 0.01). The concentrations of ADMA were higher in the cirrhotic patients with HRS than in those without this serious complication of cirrhosis. The concentration of ADMA in all the examined cirrhotic patients was higher than those obtained from healthy volunteers (1.35 +/- 0.27 micromol/L, 1.05 +/- 0.35 micromol/L and 0.76 +/- 0.21 micromol/L, respectively). In the patients with terminal alcoholic liver cirrhosis, the concentrations of ADMA and SDMA correlated with the progress of cirrhosis as well as with the development of cirrhosis complications. In the patients with HRS there was a positive correlation between creatinine and SDMA in plasma (r2 0.0756,p < 0.001) which was not found between creatinine and ADMA. The obtained results demonstrate that the increase in SDMA concentration is proportionate to the progression of chronic damage of the liver and kidneys. Increased ADMA concentration can be a causative agent of renal insufficiency in patients with cirrhosis.

A validated ultra high-pressure liquid chromatography method for separation of candesartan cilexetil impurities and its degradents in drug product

Introduction: A selective, specific, and sensitive “Ultra High-Pressure Liquid Chromatography” (UPLC) method was developed for determination of candesartan cilexetil impurities as well asits degradent in tablet formulation. Materials and Methods: The chromatographic separation was performed on Waters Acquity UPLC system and BEH Shield RP18 column using gradient elution of mobile phase A and B. 0.01M phosphate buffer adjusted pH3.0 with Orthophosphoric acid was used as mobile phase A and 95% acetonitrile with 5% Milli Q Water was used as mobile phase B. Ultraviolet (UV) detection was performed at 254nm and 210nm, where (CDS-6), (CDS-5), (CDS-7), (Ethyl Candesartan), (Desethyl CCX), (N-Ethyl), (CCX-1), (1N Ethyl Oxo CCX), (2N Ethyl Oxo CCX), (2N Ethyl) and any unknown impurity were monitored at 254nm wavelength, and two process-related impurities, trityl alcohol and MTE impurity, were estimated at 210nm. Candesartan cilexetil andimpurities were chromatographed with a total run time of 20min. Results: Calibration showed that the response of impurity was a linear function of concentration over the range limit of quantification to 2 μg/mL (r2≥0.999) and the method was validated over this range for precision, intermediate precision, accuracy, linearity, and specificity. For the precision study, percentage relative standard deviation of each impurity was <15% (n=6). Conclusion: The method was found to be precise, accurate, linear, and specific. The proposed method was successfully employed for estimation of candesartan cilexetil impurities in pharmaceutical preparations.

Ceftazidime determination in serum by high-pressure liquid chromatography.

A rapid and sensitive high-pressure liquid chromatographic method with simple sample preparation was developed for the quantitative analysis of the beta-lactam antibiotic ceftazidime (CAS 78439-06-2, Fortum). A good linear relationship was established between the peak area and the amount of ceftazidime injected over a concentration range of 1 to 200 microg/ml. The detection limit of the method was calculated to be 0.9 microg/ml. Stability was shown at 4 degrees C and at -196 degrees C for time periods of 2 h and 84 days, respectively.

PREPARATION AND CHARACTERIZATION OF LIPOSOMES CONTAINING METHANOL EXTRACT OF AERIAL PARTS OF PLATYCLADUS ORIENTALIS (L.) FRANCO

Platycladus orientalis or Thuja orientalis is a native plant of Iran. Different parts of the plant are used in the treatment of various diseases such as: gout, rheumatoid arthritis, common cold, cough, bronchitis, asthma, high blood pressure and hormonal disorders like hirsutism and baldness. Also, various organs of this species have been used as appetizer. The purpose of this study was to prepare and characterize liposomal formulations that contain methanol extract of aerial parts of P. orientalis for hirsutism treatment. Material and Methods: Plant's leaves were dried in room temperature, and powdered by grinding. Then, methanol extract was prepared by maceration method. Liposomes containing mathanol extract were produced by two methods of fusion and solvent evaporation. To evaluate mathanol extract and encapsulation efficiency of liposomes, quercetin was chosen as standard. The amount of quercetin in samples was determined by high pressure liquid chromatography (HPLC) method. Results: Mean size of liposomes prepared by solvent evaporation and fusion methods was 373 and 320 nm, respectively. According to the quercetin concentration, encapsulation efficiency of liposomes containing menthanol extract was 69.3±3.1% for solvent evaporation and 62.2±4.9% for fusion method. Conclusion: In the current study, a suitable liposomal formulation was prepared. The pharmacological activity of these carriers should be evaluated in the future study.

High pressure liquid chromatographic method for routine analysis of polycyclic aromatic hydrocarbons in airborne particulates

High pressure liquid chromatographic method for routine analysis of polycyclic aromatic hydrocarbons in airborne particulates

A validated Ultra High Pressure Liquid Chromatographic method for the characterisation of confiscated illegal slimming products containing anorexics

A fully validated UHPLC–DAD method for the identification and quantification of pharmaceutical preparations, containing molecules frequently found in illegal slimming products (sibutramine, modafinil, ephedrine, nor-ephedrine, metformin, theophyllin, caffeine, diethylpropion and orlistat) was developed. The proposed method uses a Vision HT C18-B column (2 mm × 100 mm, 1.5 μm) with a gradient using an ammonium acetate buffer pH 5.0 as aqueous phase and acetonitrile as organic modifier. The obtained method was fully validated based on its measurement uncertainty (accuracy profile). Calibration lines for all components were linear within the studied ranges. The relative bias and the relative standard deviations for all components were respectively smaller than 3.0% and 1.5%, the β-expectation tolerance limits did not exceed the acceptance limits of 10% and the relative expanded uncertainties were smaller than 3% for all of the considered components. A UHPLC–DAD method was obtained for the identification and quantification of these kind of pharmaceutical preparations, which will significantly reduce analysis times and workload for the laboratories charged with the quality control of these preparations and which can, if necessary, be coupled to a MS-detector for a more thorough characterisation.

Biochemical Measurement of Neonatal Hypoxia

Neonatal hypoxia ischemia is characterized by inadequate blood perfusion of a tissue or a systemic lack of oxygen. This condition is thought to cause/exacerbate well documented neonatal disorders including neurological impairment. Decreased adenosine triphosphate production occurs due to a lack of oxidative phosphorylation. To compensate for this energy deprived state molecules containing high energy phosphate bonds are degraded. This leads to increased levels of adenosine which is subsequently degraded to inosine, hypoxanthine, xanthine, and finally to uric acid. The final two steps in this degradation process are performed by xanthine oxidoreductase. This enzyme exists in the form of xanthine dehydrogenase under normoxic conditions but is converted to xanthine oxidase (XO) under hypoxia-reperfusion circumstances. Unlike xanthine dehydrogenase, XO generates hydrogen peroxide as a byproduct of purine degradation. This hydrogen peroxide in combination with other reactive oxygen species (ROS) produced during hypoxia, oxidizes uric acid to form allantoin and reacts with lipid membranes to generate malondialdehyde (MDA). Most mammals, humans exempted, possess the enzyme uricase, which converts uric acid to allantoin. In humans, however, allantoin can only be formed by ROS-mediated oxidation of uric acid. Because of this, allantoin is considered to be a marker of oxidative stress in humans, but not in the mammals that have uricase. We describe methods employing high pressure liquid chromatography (HPLC) and gas chromatography mass spectrometry (GCMS) to measure biochemical markers of neonatal hypoxia ischemia. Human blood is used for most tests. Animal blood may also be used while recognizing the potential for uricase-generated allantoin. Purine metabolites were linked to hypoxia as early as 1963 and the reliability of hypoxanthine, xanthine, and uric acid as biochemical indicators of neonatal hypoxia was validated by several investigators. The HPLC method used for the quantification of purine compounds is fast, reliable, and reproducible. The GC/MS method used for the quantification of allantoin, a relatively new marker of oxidative stress, was adapted from Gruber et al. This method avoids certain artifacts and requires low volumes of sample. Methods used for synthesis of MMDA were described elsewhere. GC/MS based quantification of MDA was adapted from Paroni et al. and Cighetti et al. Xanthine oxidase activity was measured by HPLC by quantifying the conversion of pterin to isoxanthopterin. This approach proved to be sufficiently sensitive and reproducible.

A fast ultra high pressure liquid chromatographic method for qualification and quantification of pharmaceutical combination preparations containing paracetamol, acetyl salicylic acid and/or antihistaminics

A fully validated UHPLC method for the identification and quantification of pharmaceutical preparations, containing paracetamol and/or acetyl salicylic acid, combined with anti-histaminics (phenylephrine, pheniramine maleate, diphenhydramine, promethazine) and/or other additives as quinine sulphate, caffeine or codein phosphate, was developed. The proposed method uses a Waters Acquity BEH C18 column (2 mm × 100 mm, 1.7 μm) with a gradient using an ammonium acetate buffer pH 4.0 as aqueous phase and methanol as organic modifier. The obtained method was fully validated based on its measurement uncertainty (accuracy profile) and robustness tests. Calibration lines for all components were linear within the studied ranges. The relative bias and the relative standard deviations for all components were respectively smaller than 1.5% and 2%, the β-expectation tolerance limits did not exceed the acceptance limits of 10% and the relative expanded uncertainties were smaller than 5% for all of the considered components. A UHPLC method was obtained for the identification and quantification of these kind of pharmaceutical preparations, which will significantly reduce analysis times and workload for the laboratories charged with the quality control of these preparations.

Development and Validation of an HPLC Method for the Determination of Thiamine and Riboflavin in Green Leafy Vegetables Using Clara‐Diastase

There is no literature on the use of the currently available form of clara-diastase (≥ 35 U/mg; Fluka Product No. 86959) in the analysis of vitamins B(1) and B(2) . Consequently, a method for the determination of total riboflavin and total thiamine in green leafy vegetables by high-pressure liquid chromatography (HPLC) following sample preparation by sequential acid hydrolysis, enzymatic hydrolysis with clara-diastase ≥ 35 U/mg, and for B1, derivatization to thiochrome, was developed and validated. Prepared samples are run at 35 °C on a 250 × 4.6 mm column of 5 μm Kromasil 100 C(18) , using a 0.8 mL/min flow of a 72:28 mixture of 5 mmol/L ammonium acetate and methanol as mobile phase and fluorescence detection. The linear response ranges and limits of detection are, respectively, 3.0 to 60.0 and 0.15 ng/mL for B(1) , and 3.1 to 155.0 and 0.103 ng/mL for B(2) . Of the 4 green leafy vegetables examined, young turnip tops had the highest B(1) and B(2) levels (0.19 and 0.20 mg/100 g fresh weight), followed by spinach (0.12 and 0.09 mg/100 g), lettuce (0.11 and 0.05 mg/100 g), and chard (0.04 mg/100 g).

From Csaba Horváth to Quality by Design: Visualizing Design Space in Selectivity Exploration of HPLC Separations

The present paper starts by taking a look back at some of the pioneering work in high pressure liquid chromatography (HPLC) that went on in Csaba Horvath’s laboratory in 1970s, through the eyes of I. Molnar. It then goes on to describe a very modern approach to HPLC method development within the Quality by Design framework: the multifactorial optimization of three critical HPLC method parameters, i.e. gradient time (t G), temperature (T), and ternary composition (B1:B2) based on 12 experiments. The effect of these experimental variables on critical resolution and selectivity was carried out in such a way as to systematically vary all three factors simultaneously.

The effect of buffering on the stability of reconstituted benzylpenicillin injection

Abstract The stability of benzylpenicillin sodium injection after reconstitution was investigated using a stability-indicating high pressure liquid chromatography (HPLC) method. Benzylpenicillin reconstituted in water for injections, diluted in 0.9 per cent sodium chloride infusion in Minibags and stored at 5C was stable for two days (based on t/90 per cent calculations). The effect of reconstituting benzylpenicillin in citrate buffer at pHs between 6 and 8 was investigated. Maximum stability was found to occur in the pH range 6.5-7.5. The stability of buffer-reconstituted benzylpenicillin diluted in 0.9 per cent sodium chloride infusion was then investigated. Results indicated that benzylpenicillin injection 600mg reconstituted in 3.6ml citrate buffer within the pH range 6.5 to 7.5, diluted in 50ml 0.9 per cent sodium chloride in Minibags and stored at 5C, may be assigned a shelf life of at least 28 days.

Evaluation of Spectrophotometric and HPLC Methods for Shikimic Acid Determination in Plants: Models in Glyphosate-Resistant and -Susceptible Crops

Endogenous shikimic acid determinations are routinely used to assess the efficacy of glyphosate in plants. Numerous analytical methods exist in the public domain for the detection of shikimic acid, yet the most commonly cited comprise spectrophotometric and high-pressure liquid chromatography (HPLC) methods. This paper compares an HPLC and two spectrophotometric methods (Spec 1 and Spec 2) and assesses the effectiveness in the detection of shikimic acid in the tissues of glyphosate-treated plants. Furthermore, the study evaluates the versatility of two acid-based shikimic acid extraction methods and assesses the longevity of plant extract samples under different storage conditions. Finally, Spec 1 and Spec 2 are further characterized with respect to (1) the capacity to discern between shikimic acid and chemically related alicyclic hydroxy acids, (2) the stability of the chromophore (t1/2), (3) the detection limits, and (4) the cost and simplicity of undertaking the analytical procedure. Overall, spectrophotometric methods were more cost-effective and simpler to execute yet provided a narrower detection limit compared to HPLC. All three methods were specific to shikimic acid and detected the compound in the tissues of glyphosate-susceptible crops, increasing exponentially in concentration within 24 h of glyphosate application and plateauing at approximately 72 h. Spec 1 estimated more shikimic acid in identical plant extract samples compared to Spec 2 and, likewise, HPLC detection was more effective than spectrophotometric determinations. Given the unprecedented global adoption of glyphosate-resistant crops and concomitant use of glyphosate, an effective and accurate assessment of glyphosate efficacy is important. Endogenous shikimic acid determinations are instrumental in corroborating the efficacy of glyphosate and therefore have numerous applications in herbicide research and related areas of science as well as resolving many commercial issues as a consequence of glyphosate utilization.

Pharmacokinetics and biodistribution of lonidamine/paclitaxel loaded, EGFR-targeted nanoparticles in an orthotopic animal model of multi-drug resistant breast cancer

The aim of this study was to assess the biodistribution and pharmacokinetics of epidermal growth factor receptor (EGFR)–targeted polymer-blend nanoparticles loaded with the anticancer drugs lonidamine and paclitaxel. Plasma, tumor, and tissue distribution profiles were quantified in an orthotopic animal model of multidrug-resistant breast cancer and were compared to treatment with nontargeted nanoparticles and to treatment with drug solution. A poly(d,l-lactide-co-glycolide)–poly(ethylene glycol)–EGFR targeting peptide (PLGA-PEG-EFGR peptide) construct was synthesized for incorporation in poly(ɛ-caprolactone) particles to achieve active EGFR targeting. An isocratic high-pressure liquid chromatography method was developed to quantify lonidamine and paclitaxel in mice plasma, tumors, and vital organs. The targeted nanoparticles demonstrated a superior pharmacokinetic profile relative to drug solution and nontargeted nanoparticles, particularly for lonidamine delivery. The first target site of accumulation was the liver, followed by the kidneys, and then the tumor mass; maximal tumor accumulation occured at 3 hours after administration. Lonidamine-paclitaxel combination therapy administered via EGFR-targeted polymer-blend nanocarriers may become a viable platform for the future treatment of multidrug-resistant cancer. From the Clinical EditorIn this study the biodistribution and pharmacokinetics of epidermal growth factor receptor (EGFR)–targeted polymer-blend nanoparticles loaded with lonidamine and paclitaxel were assessed. The targeted nanoparticles demonstrated a superior pharmacokinetic profile relative to drug solution and nontargeted nanoparticles, paving the way to new therapeutic approaches for multidrug-resistant malignancies.

Antiplasmodial Properties and Bioassay-Guided Fractionation of Ethyl Acetate Extracts fromCarica papayaLeaves

We investigated the antiplasmodial properties of crude extracts from Carica papaya leaves to trace the activity through bioassay-guided fractionation. The greatest antiplasmodial activity was observed in the ethyl acetate crude extract. C. papaya showed a high selectivity for P. falciparum against CHO cells with a selectivity index of 249.25 and 185.37 in the chloroquine-sensitive D10 and chloroquine-resistant DD2 strains, respectively. Carica papaya ethyl acetate extract was subjected to bioassay-guided fractionation to ascertain the most active fraction, which was purified and identified using high-pressure liquid chromatography (HPLC) and GC-MS (Gas chromatography-Mass spectrometry) methods. Linoleic and linolenic acids identified from the ethyl acetate fraction showed IC50 of 6.88 μg/ml and 3.58 μg/ml, respectively. The study demonstrated greater antiplasmodial activity of the crude ethyl acetate extract of Carica papaya leaves with an IC50 of 2.96 ± 0.14 μg/ml when compared to the activity of the fractions and isolated compounds.

Determination of oxidative and occupational stress in palliative care workers

In previous work, we demonstrated that some occupational workers in stressful conditions can have increases in several markers of oxidative stress when compared to other workers. We investigated two antioxidant enzymes, superoxide dismutase (SOD) and catalase (CAT) activities, and malondialdehyde (MDA) concentrations, according to demographics, lifestyle and occupational parameters in palliative care unit workers, and analyzed the relationship with occupational burnout. Fifty-two palliative care unit workers and 50 gender- and aged matched healthy individuals as controls were surveyed. Spectrophotometric and high-pressure liquid chromatography methods were used for biochemical determinations. No significant variation with respect to gender were detected with respect to SOD and CAT activities, MDA concentrations or occupational burnout. MDA concentrations increased with age in controls and palliative care unit workers, and we observed significant differences in MDA between controls and palliative care unit workers for all age groups. Significant variation in MDA concentrations were detected between unmarried (287.22±8.31 nmol/mg hemoglobin) and married individuals (317.18±6.24 nmol/mg hemoglobin), but not with respect to divorced individuals (288.41±5.64 nmol/mg hemoglobin). Significant differences were detected between smokers and non-smokers for SOD, CAT and MDA, but not for alcohol, coffee, tea or cola consumption. Significant differences were seen in MDA concentrations between those who frequently practice some kind of sport (280.59±7.62 nmol/mg hemoglobin) and those who never practice any kind of sport (299.12±8.09 nmol/mg hemoglobin), and between those who frequently ate fruit and greens (291.05±8.11 nmol/mg hemoglobin) and those who never eat fruit and greens (316.31±7.42 nmol/mg hemoglobin). SOD activity and MDA concentrations are higher in palliative care workers who work the evening and night shifts (p<0.01), and these workers also show significantly higher levels of stress. Our findings suggest that oxidative stress, occupational stress and occupational burnout levels are similar in men and women. Occupational stress increases oxidative stress levels probably as a response to increased generation of reactive oxygen species. Working during the evening and night shifts increases oxidative levels and burnout levels.

Inclusion complexes of fluconazole with β-cyclodextrin: physicochemical characterization and in vitro evaluation of its formulation

Fluconazole (FZ) is a triazole antifungal drug administered orally or intravenously. It is employed for the treatment of mycotic infections. However, the efficacy of FZ is limited with its poor aqueous solubility and low dissolution rate. One of the important pharmaceutical advantages of cyclodextrins is to improve pharmacological efficacy of drugs due to increasing their aqueous solubility. The aim of present study was to prepare an inclusion complex of FZ and β-cyclodextrin (β-CD) to improve the physicochemical and biopharmaceutical properties of FZ. The effects of β-CD on the solubility of FZ were investigated according to the phase solubility technique. Complexes were prepared with 1:1 M ratio by different methods namely, freeze-drying, spray-drying, co-evaporation and kneading. For the characterization of FZ/β-CD complex, FZ amount, practical yield %, thermal, aqueous solubility, XRD, FT-IR and NMR (1H and 13C) analysis were performed. In vitro dissolution from hard cellulose capsules containing FZ/β-CD complexes was compared to pure FZ and its commercial capsules and evaluated by f1 (difference) and f2 (similarity) factors. Paddle method defined in USP 31 together with high pressure liquid chromatographic method were used in in vitro dissolution experiments. It was found that solubility enhancement by FZ/β-CD complexes depends on the type of the preparation method. High release of active agent from hard cellulose capsules prepared with β-CD complexes compared to commercial capsules was attributed to the interactions between β-CD and active agent, high energetic amorphous state and inclusion complex formation.