Development and Comparative Evaluation of Two Different Label-Free and Sensitive Fluorescence Platforms for Analysis of Olaparib: A Recently FDA-Approved Drug for the Treatment of Ovarian and Breast Cancer.

Olaparib (OLA) is a PARP inhibitor drug which has been recently approved by the Food and Drug Administration (FDA) for the treatment of ovarian and breast cancer. A convenient analytical tool for the quantitation of OLA in its dosage form and plasma samples was urgently needed. This study describes, for the first time, the development of two different label-free and sensitive fluorescence-based platforms for the pharmaceutical and bioanalysis of OLA. These platforms were microwell-assisted with a fluorescence microplate reader (MW-FLR) and high-performance liquid chromatography with fluorescence detection (HPLC-FD). Both MW-FLR and HPLC-FD employed the native fluorescence of OLA as an analytical signal. The MW-FLR involved measuring the fluorescence signals in 96-well white-opaque plates. The HPLC-FD involved chromatographic separation of OLA and duvelisib (DUV), as an internal standard on a Nucleosil-CN HPLC column (250 mm length × 4.6 mm i.d., 5 µm particle diameter) with a mobile phase composed of acetonitrile: water (25:75, v/v) pumped at a flow rate of 1.7 mL/min. Elution of OLA and DUV was detected using a fluorescence detector. The optimal conditions of both MW-FLR and HPLC-FD were established, and they were validated according to the guidelines of the International Council for Harmonization for the validation of analytical procedures. The linear ranges of MW-FLR and HPLC-FD were 25-1000 and 5-200 ng/mL, respectively, with limits of detection of 15 and 1.7 ng/mL, respectively. The accuracy and precision of both platforms were confirmed as the recovery values were ≥98.2% and the relative standard deviations (RSD) were ≤2.89%. Both methodologies were satisfactorily applied to the quantitation of OLA in its commercial dosage form (Lynparza® tablets) and plasma samples with high accuracy and precision. The greenness of both MW-FLR and HPLC-FD was assessed using two different multiple parameter-based metric tools, and the results proved their greenness and adherence to the requirements of green analytical approaches. Both platforms have simple procedures and acceptable levels of analytical throughput. In conclusion, the proposed MW-FLR and HPLC-FD are valuable tools for routine use in quality control and clinical laboratories for the quantitation of OLA for the purposes of pharmaceutical quality control, pharmacokinetic studies, and bioequivalence testing.

Synthesis, spectroscopic and computational characterization of charge transfer complex of remdesivir with chloranilic acid: Application to development of novel 96-microwell spectrophotometric assay.

Remdesivir (REM) is an adenosine triphosphate analog antiviral drug that has received authorization from European Commission and approval from the U.S. Food and Drug Administration for treatment of coronavirus disease 2019 (Covid-19). This study, describes, for the first time, the synthesis of a novel charge transfer complex (CTC) between REM, as electron donor, with chloranilic acid (CLA), as π electron acceptor. The CTC was characterized using different spectroscopic and thermogravimetric techniques. UV-visible spectroscopy ascertained the formation of the CTC in methanol via formation of a new broad absorption band with maximum absorption peak (λmax) at 530 nm. The molar absorptivity (ε) of the complex was 3.33 × 103 L mol-1 cm-1 and its band gap energy was 1.91 eV. The stoichiometric ratio of REM:CLA was found to be 1:1. The association constant of the complex was 1.11 × 109 L mol-1, and its standard free energy was 5.16 × 104 J mole-1. Computational calculation for atomic charges of energy minimized REM was conducted, the site of interaction on REM molecule was assigned and the mechanism of the reaction was postulated. The solid-state CTC was further characterized by FT-IR and 1H NMR spectroscopic techniques. Both FT-IR and 1H NMR confirmed the formation of the CTC and its structure. The reaction was adopted as a basis for developing a novel 96-microwell spectrophotometric method (MW-SPA) for REM. The assay limits of detection and quantitation were 3.57 and 10.83 µg/well, respectively. The assay was validated, and all validation parameters were acceptable. The assay was implemented successfully with great precision and accuracy to the determination of REM in its bulk form and pharmaceutical formulation (injection). This assay is simple, economic, and more importantly, has high throughput property. Therefore, the assay can be valuable for routine in quality control laboratories for analysis of REM's bulk form and pharmaceutical injection.

Experimental and Computational Evaluation of Chloranilic Acid as an Universal Chromogenic Reagent for the Development of a Novel 96-Microwell Spectrophotometric Assay for Tyrosine Kinase Inhibitors.

The tyrosine kinase inhibitors (TKIs) are chemotherapeutic drugs used for the targeted therapy of various types of cancer. This work discusses the experimental and computational evaluation of chloranilic acid (CLA) as a universal chromogenic reagent for developing a novel 96-microwell spectrophotometric assay (MW-SPA) for TKIs. The reaction resulted in an instantaneous formation of intensely purple colored products with TKIs. Spectrophotometric results confirmed that the reactions proceeded via the formation of charge-transfer complexes (CTCs). The physical parameters were determined for the CTCs of all TKIs. Computational calculations and molecular modelling for the CTCs were conducted, and the site(s) of interaction on each TKI molecule were determined. Under the optimized conditions, Beer's law correlating the absorbances of the CTCs with the concentrations of TKIs were obeyed in the range of 10-500 µg/well with good correlation coefficients (0.9993-0.9998). The proposed MW-SPA fully validated and successfully applied for the determination of all TKIs in their bulk forms and pharmaceutical formulations (tablets). The proposed MW-SPA is the first assay that can analyze all the TKIs on a single assay system without modifications in the detection wavelength. The advantages of the proposed MW-SPA are simple, economic and, more importantly, have high throughput.

Modulating sphingosine 1-phosphate signaling with DOP or FTY720 alleviates vascular and immune defects in mouse sepsis.

Sepsis is a systemic inflammatory response to pathogens and a leading cause of hospital related mortality worldwide. Sphingosine 1-phosphate (S1P) regulates multiple cellular processes potentially involved in the pathogenesis of sepsis, including antigen presentation, lymphocyte egress, and maintenance of vascular integrity. We thus explored the impact of manipulating S1P signaling in experimental polymicrobial sepsis in mice. Administration of 4-deoxypyridoxine (DOP), an inhibitor of the S1P-degrading enzyme S1P-lyase, or of the sphingosine analog FTY720 that serves as an S1P receptor agonist after phosphorylation ameliorated morbidity, improved recovery from sepsis in surviving mice, and reduced sepsis-elicited hypothermia and body weight loss. Treated mice developed lymphopenia, leading to an accumulation of lymphocytes in peripheral lymph nodes, and reduced bacterial burden in liver, but not in blood. Sepsis-induced upregulation of mRNA expression of cytokines in spleen remained unchanged, but reduction of IL-6, TNF-α, MCP-1, and IL-10 in plasma was evident. DOP and FTY720 treatment significantly reduced levels of Evans blue leakage from blood into liver and lung, decreased hematocrit values, and lowered plasma levels of VEGF-A in septic mice. Collectively, our results indicate that modulation of S1P signaling showed a protective phenotype in experimental sepsis by modulating vascular and immune functions.

Simplified risk stratification criteria for identification of patients with MRSA bacteremia at low risk of infective endocarditis: implications for avoiding routine transesophageal echocardiography in MRSA bacteremia.

The aim of this study was to identify patients with methicillin-resistant Staphylococcus aureus (MRSA) bacteremia with low risk of infective endocarditis (IE) who might not require routine trans-esophageal echocardiography (TEE). We retrospectively evaluated 398 patients presenting with MRSA bacteremia for the presence of the following clinical criteria: intravenous drug abuse (IVDA), long-term catheter, prolonged bacteremia, intra-cardiac device, prosthetic valve, hemodialysis dependency, vertebral/nonvertebral osteomyelitis, cardio-structural abnormality. IE was diagnosed using the modified Duke criteria. Of 398 patients with MRSA bacteremia, 26.4 % of cases were community-acquired, 56.3 % were health-care-associated, and 17.3 % were hospital-acquired. Of the group, 44 patients had definite IE, 119 had possible IE, and 235 had a rejected diagnosis. Out of 398 patients, 231 were evaluated with transthoracic echocardiography (TTE) or TEE. All 44 patients with definite IE fulfilled at least one criterion (sensitivity 100 %). Finally, a receiver operator characteristic (ROC) curve was obtained to evaluate the total risk score of our proposed criteria as a predictor of the presence of IE, and this was compared to the ROC curve of a previously proposed criteria. The area under the ROC curve for our criteria was 0.710, while the area under the ROC curve for the criteria previously proposed was 0.537 (p < 0.001). The p-value for comparing those 2 areas was less than 0.001, indicating statistical significance. Patients with MRSA bacteremia without any of our proposed clinical criteria have very low risk of developing IE and may not require routine TEE.

Modulation of GLO1 Expression Affects Malignant Properties of Cells.

The energy metabolism of most tumor cells relies on aerobic glycolysis (Warburg effect) characterized by an increased glycolytic flux that is accompanied by the increased formation of the cytotoxic metabolite methylglyoxal (MGO). Consequently, the rate of detoxification of this reactive glycolytic byproduct needs to be increased in order to prevent deleterious effects to the cells. This is brought about by an increased expression of glyoxalase 1 (GLO1) that is the rate-limiting enzyme of the MGO-detoxifying glyoxalase system. Here, we overexpressed GLO1 in HEK 293 cells and silenced it in MCF-7 cells using shRNA. Tumor-related properties of wild type and transformed cells were compared and key glycolytic enzyme activities assessed. Furthermore, the cells were subjected to hypoxic conditions to analyze the impact on cell proliferation and enzyme activities. Our results demonstrate that knockdown of GLO1 in the cancer cells significantly reduced tumor-associated properties such as migration and proliferation, whereas no functional alterations where found by overexpression of GLO1 in HEK 293 cells. In contrast, hypoxia caused inhibition of cell growth of all cells except of those overexpressing GLO1. Altogether, we conclude that GLO1 on one hand is crucial to maintaining tumor characteristics of malignant cells, and, on the other hand, supports malignant transformation of cells in a hypoxic environment when overexpressed.

A validated high-throughput UHPLC-MS/MS assay for accurate determination of rivaroxaban in plasma sample.

Rivaroxaban is a novel, selective and potent oral direct factor Xa inhibitor, therapeutically indicated in the treatment of thromboembolic diseases. Like traditional anticoagulants, routine coagulation monitoring of rivaroxaban is not necessary, but important in some clinical circumstances. In this study, a sensitive UHPLC-MS/MS assay for rapid determination of rivaroxaban in human plasma was developed and validated. Rivaroxaban and its internal standard (IS) were extracted from plasma using acetonitrile as protein precipitating agent. An isocratic mobile phase of acetonitrile: 10 mM ammonium acetate (80:20, v/v) at a flow rate of 0.3 mL/min was used for the separation of rivaroxaban and IS. Both rivaroxaban and IS was eluted within 1 min with a total run time of 1.5 min only. Electrospray ionization source in positive mode was used for the detections of rivaroxaban and IS. Precursor to product ion transition of m/z 436.00 > 144.87 for rivaroxaban and m/z 411.18 > 191.07 for IS were used in multiple reaction monitoring mode. Developed assay was fully validated in terms of selectivity, linearity, accuracy, precision, recovery, matrix effects and stability using official guideline on bioanalytical method.

Photosensitivity of gallium-doped silica core fiber to 193 nm ArF excimer laser.

Grating inscription in a Ga-doped silica core fiber (~5 wt. % Ga) has been demonstrated using ArF (193 nm) and KrF (248 nm) excimer lasers. In a comparative study with germanosilicate fiber with similar Ge concentration, a Ga-doped silica core fiber shows greater photosensitivity to an ArF excimer laser due to the higher absorbance in the region of 190-195 nm. In addition, the photosensitivity of a Ga-doped silica core fiber has been greatly enhanced with hydrogenation. Ga-doped fibers are potential photosensitive fibers for fiber Bragg grating production with an ArF excimer laser.

Charge-transfer reaction of 2,3-dichloro-1,4-naphthoquinone with crizotinib: Spectrophotometric study, computational molecular modeling and use in development of microwell assay for crizotinib.

The reaction of 2,3-dichloro-1,4-naphthoquinone (DCNQ) with crizotinib (CZT; a novel drug used for treatment of non-small cell lung cancer) was investigated in different solvents of varying dielectric constants and polarity indexes. The reaction produced a red-colored product. Spectrophotometric investigations confirmed that the reaction proceeded through charge-transfer (CT) complex formation. The molar absorptivity of the complex was found to be linearly correlated with the dielectric constant and polarity index of the solvent; the correlation coefficients were 0.9567 and 0.9069, respectively. The stoichiometric ratio of DCNQ:CZT was found to be 2:1 and the association constant of the complex was found to be 1.07 × 10(2) l/mol. The kinetics of the reaction was studied; the order of the reaction, rate and rate constant were determined. Computational molecular modeling for the complex between DCNQ and CZT was conducted, the sites of interaction on CZT molecule were determined, and the mechanism of the reaction was postulated. The reaction was employed as a basis in the development of a novel 96-microwell assay for CZT in a linear range of 4-500 µg/ml. The assay limits of detection and quantitation were 2.06 and 6.23 µg/ml, respectively. The assay was validated as per the guidelines of the International Conference on Harmonization (ICH) and successfully applied to the analysis of CZT in its bulk and capsules with good accuracy and precision. The assay has high throughput and consumes a minimum volume of organic solvents thus it reduces the exposures of the analysts to the toxic effects of organic solvents, and significantly reduces the analysis cost.

Exploring glyoxalase 1 expression in prostate cancer tissues: targeting the enzyme by ethyl pyruvate defangs some malignancy-associated properties.

BACKGROUND: The glyoxalase (GLO)1 is part of a ubiquitous detoxification system in the glycolytic pathway of normal and tumor cells. It protects against cellular damage caused by cytotoxic metabolites. METHODS: Aiming at exploring the role of GLO1 in prostate cancer, we evaluated and targeted the expression of GLO1 in prostate cancer tissues and cell lines and analyzed its correlation with grading systems and tumor growth indices. RESULTS: Immunohistochemical studies on 37 prostate cancer specimens revealed a positive correlation between Helpap-grading and the cytoplasmic (P = 0.002)/nuclear (P = 0.006) GLO1 level. A positive correlation between Ki-67 proliferation marker and the cytoplasmic GLO1 (P = 0.006) was evident. Furthermore, the highest GLO1 level was detected in the androgen-sensitive LNCaP compared to the androgen-independent Du-145 and PC-3 prostate cell lines and the breast cancer cell MCF-7, both at protein and mRNA level. Treating cancer cells with ethyl pyruvate was found to defang some malignancy-associated properties of cancer cells including proliferation, invasion and anchorage-independent growth. In vitro results revealed that the potency of ethyl pyruvate is increased when cells are metabolically activated by growth stimulators, for example, by fetal calf serum, dihydrotestosterone, tumor growth factor-ß1 and leptin. CONCLUSIONS: The positive correlation of GLO1 expression level in prostate cancer tissues with the pathological grade and proliferation rate may assign GLO1 as a risk factor for prostate cancer development and progression. Furthermore, our data indicate that inhibitors of GLO1 might be useful to decelerate the cancer cell growth by a novel therapeutic approach that we may call "induced metabolic catastrophe."

Sphingosine 1-phosphate in blood: function, metabolism, and fate.

Sphingosine 1-phosphate (S1P) is a lipid metabolite and a ligand of five G protein-coupled cell surface receptors S1PR1 to S1PR5. These receptors are expressed on various cells and cell types of the immune, cardiovascular, respiratory, hepatic, reproductive, and neurologic systems, and S1P has an impact on many different pathophysiological conditions including autoimmune, cardiovascular, and neurodegenerative diseases, cancer, deafness, osteogenesis, and reproduction. While these diverse signalling properties of S1P have been extensively reviewed, the particular role of S1P in blood is still a matter of debate. Blood contains the highest S1P concentration of all body compartments, and several questions are still not sufficiently answered: Where does it come from and how is it metabolized? Why is the concentration of S1P in blood so high? Are minor changes of the high blood S1P concentrations physiologically relevant? Do blood cells and vascular endothelial cells that are constantly exposed to high blood S1P levels still respond to S1P via S1P receptors? Recent data reveal new insights into the functional role and the metabolic fate of blood-borne S1P. This review aims to summarize our current knowledge regarding the source, secretion, transportation, function, metabolism, and fate of S1P in blood.

The role of T helper (TH)17 cells as a double-edged sword in the interplay of infection and autoimmunity with a focus on xenobiotic-induced immunomodulation.

Extensive research in recent years suggests that exposure to xenobiotic stimuli plays a critical role in autoimmunity induction and severity and that the resulting response would be exacerbated in individuals with an infection-aroused immune system. In this context, heavy metals constitute a prominent category of xenobiotic substances, known to alter divergent immune cell responses in accidentally and occupationally exposed individuals, thereby increasing the susceptibility to autoimmunity and cancer, especially when accompanied by inflammation-triggered persistent sensitization. This perception is learned from experimental models of infection and epidemiologic studies and clearly underscores the interplay of exposure to such immunomodulatory elements with pre- or postexposure infectious events. Further, the TH17 cell subset, known to be associated with a growing list of autoimmune manifestations, may be the "superstar" at the interface of xenobiotic exposure and autoimmunity. In this review, the most recently established links to this nomination are short-listed to create a framework to better understand new insights into TH17's contributions to autoimmunity.

Induction of osteoporosis with its influence on osteoporotic determinants and their interrelationships in rats by DEXA.

BACKGROUND: As women are the population most affected by multifactorial osteoporosis, research is focused on unraveling the underlying mechanism of osteoporosis induction in rats by combining ovariectomy (OVX) either with calcium, phosphorus, vitamin C and vitamin D2/D3 deficiency, or by administration of glucocorticoid (dexamethasone). MATERIAL/METHODS: Different skeletal sites of sham, OVX-Diet and OVX-Steroid rats were analyzed by Dual Energy X-ray Absorptiometry (DEXA) at varied time points of 0, 4 and 12 weeks to determine and compare the osteoporotic factors such as bone mineral density (BMD), bone mineral content (BMC), area, body weight and percent fat among different groups and time points. Comparative analysis and interrelationships among osteoporotic determinants by regression analysis were also determined. RESULTS: T scores were below-2.5 in OVX-Diet rats at 4 and 12 weeks post-OVX. OVX-diet rats revealed pronounced osteoporotic status with reduced BMD and BMC than the steroid counterparts, with the spine and pelvis as the most affected skeletal sites. Increase in percent fat was observed irrespective of the osteoporosis inducers applied. Comparative analysis and interrelationships between osteoporotic determinants that are rarely studied in animals indicate the necessity to analyze BMC and area along with BMD in obtaining meaningful information leading to proper prediction of probability of osteoporotic fractures. CONCLUSIONS: Enhanced osteoporotic effect observed in OVX-Diet rats indicates that estrogen dysregulation combined with diet treatment induces and enhances osteoporosis with time when compared to the steroid group. Comparative and regression analysis indicates the need to determine BMC along with BMD and area in osteoporotic determination.

Vinpocetine (A comprehensive profile).

Vinpocetine (VIN) is a herbal supplement extracted from the periwinkle plant. It is a multi-action agent, which is used to treat various neurological disorders such as Alzheimer's and Parkinson's disease. Vinpocetine has also anti-inflammatory, analgesic, antioxidant property and treats various thinking and memory problems. Currently, vinpocetine is also available in the market as a dietary supplement to enhance cognition and memory. This profile explains the physicochemical properties, methods of preparation, content of related impurities and different spectroscopical behavior of vinpocetine. It also discusses the reported methods of analysis of the drug, which include Compendial Methods, Electrochemical Methods, Spectrophotometric Methods and Chromatographic Methods of analysis. Furthermore, this profile explains the stability of the drug subjected to stress conditions of acid, alkaline and photolytic degradation. In addition, the clinical applications of the drug, its uses, side effects, dosing information, pharmacokinetics and mechanism of action are also discussed.

Synthesis, spectroscopic and computational studies on hydrogen bonded charge transfer complex of duvelisib with chloranilic acid: Application to development of novel 96-microwell spectrophotometric assay.

Duvelisib (DUV) is a is a small-molecule with inhibitory action for phosphoinositide 3-kinase (PI3K). It has been recently approved for the effective treatment of chronic lymphocytic leukemia (CLL) and small lymphocytic lymphoma (SLL). Novel charge transfer complex (CTC) between DUV, as electron donor, with chloranilic acid (CLA), as π electron acceptor has been synthesized and characterized using different spectroscopic and thermogravimetric techniques. UV-visible spectroscopy ascertained the formation of the CTC in different solvents of varying polarity indexes and dielectric constants via formation of new broad absorption band with maximum absorption peak (λmax) in the range of 488-532 nm. The molar absorptivity of the CTC was dependent on the polarity index and dielectric constant of the solvent; the correlation coefficients were 0.9955 and 0.9749, respectively. The stoichiometric ratio of DUV:CLA was 1:1. Electronic spectral analysis was conducted for characterization of the complex in terms of its electronic constants. Computational calculation for atomic charges of energy minimized DUV was conducted and the site of interaction on DUV molecule was assigned. The solid-state CTC of DUV:CLA (1:1) was synthesized, and its structure was characterized by UV-visible, mass, FT-IR, and 1H NMR spectroscopic techniques. Both FT-IR and 1H NMR confirmed that both CT and hydrogen bonding contributed to the molecular composition of the complex. The reaction was adopted as a basis for developing a novel 96-microwell spectrophotometric assay (MW-SPA) for DUV. The assay limits of detection and quantitation were 0.57 and 1.72 µg/well, respectively. The assay was validated and all validation parameters were acceptable. The method was implemented successfully with great precision and accuracy to the analysis of the DUV in its bulk and capsules.

Enantioselective organocatalytic Diels-Alder reactions: a density functional theory and kinetic isotope effects study.

The mechanism and enantioselectivity of the organocatalytic Diels-Alder reaction were computationally investigated by density functional theory at the B3LYP/6-31G(d) level of theory. The uncatalyzed Diels-Alder reaction was also studied to explore the effect of the organocatalyst on this reaction in terms of energetics, selectivity, and mechanism. The catalyzed reaction showed improved endo/exo selectivity, and the free energy of activation was significantly lowered in the presence of the catalyst. Both uncatalyzed and catalyzed reactions exhibited concerted asynchronous reaction mechanism with the degree of asynchronicity being more evident in the presence of the catalyst. The Corey's experimentally derived predictive selection rules for the outcome of the organocatalytic Diels-Alder reaction were also theoretically analyzed, and an excellent agreement was found between experiment and theory.

Spectrophotometric and computational investigations of charge transfer complexes of chloranilic acid with tyrosine kinase inhibitors and application to development of novel universal 96-microwell assay for their determination in pharmaceutical formulations.

The tyrosine kinase inhibitors (TKIs) are chemotherapeutic drugs used for targeted therapy of various types of cancer. In literature, there is no existing universal chromogenic reagent used for development of spectrophotometric assay for all TKIs regardless the diversity of their chemical structures. This work discusses, for the first time, the experimental and computational evaluation of chloranilic acid (CLA) as a universal chromogenic reagent for developing a novel 96-microwell spectrophotometric assay (MW-SPA) for TKIs. The reaction of CLA with seven TKIs was examined in different organic solvents of various dielectric constants and polarity indexes. The reaction resulted in an instantaneous formation of intensely purple coloured products with all the investigated TKIs. Spectrophotometric investigations confirmed that the reactions proceeded via the formation of charge-transfer complexes (CTC). The physical parameters (molar absorptivity, molar ratio, association constant and standard free energy) were determined for the CTC of all TKIs. Computational calculations for the relative electron densities on each atom of the TKI molecule and molecular modelling for the CTC were conducted, and the site(s) of interaction on each TKI molecule were determined. Under the optimized conditions, Beer's law correlating the absorbances of the CTC with the concentrations of TKIs were obeyed in the range of 5-500 µg/well with good correlation coefficients (0.9991-0.9998). The limits of detection and quantitation were in the ranges of 1.89-5.09 and 5.74-15.42 µg/well, respectively. The proposed MW-SPA showed high precisions as the values of the relative standard deviations did not exceed 2.01 and 2.45% for the intra- and inter-assay precision, respectively. The accuracy of MW-SPA was proved by recovery studies as the recovery values were in the range of 98.8-103.7%. The proposed MW-SPA was successfully applied for the determination of all TKIs in their bulk forms and pharmaceutical formulations (tablets) with good accuracy and precisions. The proposed MW-SPA is the first assay that can analyse all the TKIs on a single assay system without modifications in the detection wavelength. Additional advantages of the proposed MW-SPA are simple, economic, and more importantly have high throughput. Therefore, the assay can be helpful and beneficial for routine analysis of TKIs in their pharmaceutical formulations in quality control laboratories.

A simple and sensitive HILIC-based UHPLC-MS/MS method for quantifying of rivaroxaban in dried blood spots: Application in comparison with the plasma sample method.

Rivaroxaban, indicated for the treatment of atrial fibrillation, deep vein thrombosis, pulmonary embolism, and coronary or peripheral artery disease, is one of the most frequently used direct oral anticoagulants. Therapeutic drug monitoring [TDM] is essential to minimize bleeding and thrombosis during personalized rivaroxaban treatment. An efficient and reliable analytical technique is required to quatify the rivaroxaban during its therapeutic indication. Dried blood spots (DBSs) sampling is a convenient bioanalytical method with minimal invasive blood drawing, long-term stability, and low shipment and storage costs. Therfore, DBS sampling technique is growing rapidly for TDM of drugs in medical care. This study developed an ultra high performance liquid chromatography-tandem mass spectrometry method of quantitating rivaroxaban in DBSs samples using the isotopic labeled analog (rivaroxaban-d4) as an internal standard (IS). Rivaroxaban and IS were separated on an Acquity HILIC column and eluted with a mobile-phase composition of acetonitrile and 20 mM ammonium acetate in the ratio of 95:5 at a flow rate of 0.3 mL/min. The precursor-to-product ion transitions of 436.03 ˃ 144.9 for rivaroxaban and 440.04 ˃ 144.9 for IS were used to quantify in multiple reaction monitoring mode. The method was accurate and precise in the 2.06-1000 ng/mL calibration range without hematocrit and blood spot volume effects. Rivaroxaban was stable in DBSs samples under different anticipated storage and temperature conditions. We observed good correlation between the plasma concentration and the DBSs concentration, indicating that the proposed DBSs method is suitable for monitoring the rivaroxaban concentration using a simple and convenient sample collection procedure.

A Highly Sensitive Nonextraction-Assisted HPLC Method with Fluorescence Detection for Quantification of Duvelisib in Plasma Samples and its Application to Pharmacokinetic Study in Rats.

BACKGROUND: Duvelisib (DUV) is a new oral phosphoinositide-3-kinase (PI3K)-δ and PI3K-γ inhibitor. It has been recently granted an accelerated approval for treatment of adult patients with relapsed or refractory chronic lymphocytic leukemia (CLL) and small lymphocytic lymphoma (SLL). It is also effective in therapy of T-cell lymphoma, solid tumors, and non-Hodgkin's lymphoma. In literature, there is no method valid for quantitation of DUV in human plasma for its therapeutic monitoring and pharmacokinetic studies. PURPOSE: The purpose of this study is the establishment of a highly sensitive HPLC method with fluorescence detection for quantitation of DUV in plasma for its therapeutic monitoring and pharmacokinetic studies of DUV. METHODS: The resolution of DUV and the internal standard (IS) olaparib (OLA) was achieved on Nucleosil CN column, with a mobile phase composed of acetonitrile:water (25:75, v/v) at a flow rate of 1.7 mL min-1. The fluorescence of both DUV and OLA was detected at 410 nm after excitation at 280 nm. The method was validated according to the guidelines of bioanalytical method validation. RESULTS: The method was linear in the range of 5-100 ng mL-1, and its limit of detection (LOD) and limit of quantitation (LOQ) were 2.12 ng mL-1 and 7 ng mL-1, respectively. The precisions of the method were ≤ 8.26%, and its accuracies were ≥ 95.32%. All the other validation parameters were satisfactory. The proposed method was successfully employed to the investigation of the pharmacokinetic profile of DUV in rats following a 25 mg/kg single dose of oral administration. CONCLUSION: The method is characterized with high sensitivity, accuracy, simple sample pretreatment, rapidity, eco-friendly as it consumes low volumes of organic solvent in the mobile phase and has high analysis throughput as its run time was short (~ 10 min).

Improving the Solubilization and Bioavailability of Arbidol Hydrochloride by the Preparation of Binary and Ternary ß-Cyclodextrin Complexes with Poloxamer 188.

In the current study, the effect of poloxamer 188 on the complexation efficiency and dissolution of arbidol hydrochloride (ADL), a broad-spectrum antiviral agent, with ß-cyclodextrin (ß-CD) was investigated. Phase solubility studies confirmed a stoichiometry of a 1:1 ratio for both ADL:ß-CD and ADL/ß-CD with a 1% poloxamer 188 system with an AL type of phase solubility curve. The stability constants (K1:1) calculated from the AL type diagram were 550 M-1 and 2134 M-1 for AD:ß-CD and ADL/ß-CD with 1% poloxamer 188, respectively. The binary ADL/ß-CD and ternary ADL/ß-CD with 1% poloxamer 188 complexes were prepared by kneading and a solvent evaporation method and were characterized by aqueous solubility, FTIR, PXRD, DSC and SEM in vitro studies. The solubility (13.1 fold) and release of ADL were markedly improved in kneaded ternary ADL/ß-CD with 1% poloxamer 188 (KDB). The binding affinity of ADL and ß-CD was confirmed by 1H NMR and 2D ROSEY studies. The ternary complex (KDB) was further subjected for in vivo pharmacokinetic studies in rats and a significant improvement in the bioavailability (2.17 fold) was observed in comparison with pure ADL. Therefore, it can be concluded that the solubilization and bioavailability of ADL can be remarkably increased by ADL/ß-CD complexation in the presence of a third component, poloxamer 188.