Development of a SPE-LC-MS Method for the Quantitation of Palbociclib and Abemaciclib in Human Plasma.

Palbociclib and abemaciclib are two cyclin-dependent kinases 4 and 6 used for breast cancer treatment. Levels of these medicines present a significant interindividual variability, so monitoring those concentrations might be necessary in therapy. Most of the methods presented so far in the literature use simple protein precipitation of plasma proteins as sample preparation method followed by direct injection of the supernatant into the LC instrument, preceded or not by a simple filtration step. Within that approach, the probability of injecting proteins in the chromatographic system is increased. With the purpose of obtaining a cleaner extract of the drugs, we developed and validated a simple and accurate LC-MS method for determining palbociclib and abemaciclib in human plasma. Solid phase extraction (SPE) using Oasis PRiME HLB® cartridges was used for plasma sample preparation. The method provided clean extracts with a recovery extraction higher than 85% for both compounds. Separation was achieved by high-performance liquid chromatography (HPLC), using a C18 (4.6 × 50 mm) column, with a gradient elution of ammonium acetate/acetic acid-acetonitrile as the mobile phase. Detection was performed by mass spectrometry (MS) in single ion recording (SIR) mode. Intra-day and inter-day precision data for both analytes were 3.8-7.2% and 3.6-7.4%, respectively. Calibration curves were both linear between 2 and 400 ng/mL with a correlation coefficient higher than 0.998. The LC-MS method can be used to quantify the drugs in human plasma in routine analysis. The method proved to be useful in determining real plasma levels in patients involved in cancer therapy. Drug concentrations were determined in a 10 min run-time, including re-equilibration of the column.

Simultaneous Determination of Two Tyrosine Kinase Inhibitors in Tablets by HPLC-MS analysis.

The class of tyrosine kinase inhibitors (TKIs) is represented by a group of compounds which are currently used in the treatment of different types of cancer. These oral medicines present a narrow therapeutic index and a large inter-and intra-individual variability. Within this work, a simple, accurate and rapid reversed phase ultra-high-performance liquid chromatographic (RP-UHPLC) method with mass spectrometric (MS) detection for simultaneous analysis of two TKIs, ibrutinib and ruxolitinib, using pentoxifylline as internal standard (IS) in tablet dosage forms is presented. The separation was carried out on a Waters (Milford, Massachusetts, USA) Arc System coupled with a Waters QDa mass detector. The column used was a Waters CORTECS C18 (4.6×50mm, 2.7µm); a gradient elution was carried out using a mixture of ammonium formate 10 mM aqueous solution and acetonitrile. The flow rate of the mobile phase was set to 0.5mL/min. The column temperature was equilibrated to 40°C. The injected volume was 5µL. All samples were kept at 20°C during the entire analysis. Mass spectra were recorded in positive ionization mode in the range of m/z 100-400 for ruxolitinib and m/z 100-500 for ibrutinib. Quantification was established in single ion recording (SIR) mode for each compound, using pentoxifylline as internal standard. The method was validated according to International Guidelines in terms of stability, limit of detection, limit of quantitation, linearity, precision and accuracy. The validated method can be successfully applied for simultaneous determination of TKIs in tablet dosage forms.

Hydroxyapatite-ciprofloxacin delivery system: Synthesis, characterisation and antibacterial activity.

The main objective of this study was to synthesize hydroxyapatite-ciprofloxacin composites using a chemical precipitation method and to evaluate the properties and in vitro release profile of the drug from the hydroxyapatite-ciprofloxacin composites. Composite characterization was achieved by FT-IR, XRD and DLS. Ciprofloxacin determination was accomplished by HPLC, resulting in good incorporation efficiency of the drug (18.13 %). The in vitro release study (Higuchi model C = K t1/2 and Ritger-Peppas model, C = K t0.6) showed a diffusion-controlled mechanism. The antibacterial activity showed that the bacterial growth inhibition zones were approximately equal for the synthesis composites and for the mechanical mixture on the Staphylococcus aureus germ. The use of hydroxyapatite, which is a biocompatible, bioactive and osteoconductive material, with ciprofloxacin, which has good antibacterial activity in this composite, makes it suitable for the development of bone grafts. Furthermore, the synthesis process allows a slow local release of the drug.

Magnetic nanoparticles-based therapy for malignant mesothelioma.

This work was aimed to analyze the versatility of the chick embryo chorioallantoic membrane (CAM) as in vivo model for the study of the malignant pleural mesothelioma (MPM) and the therapeutic potential of Fe3O4÷salicylic acid magnetic nanoparticles (SaMNPs) on MPM cells. The antitumor effects of SaMNPs were studied by in vitro and in vivo tests on CARM-L12 TG3 rat malignant mesothelioma cells and human MPM xenografts implanted on CAMs. In order to assess the human MPM xenograft growth characteristics, calretinin, HBME-1 (Hector Battifora mesothelial epitope-1), and cytokeratins immunohistochemical stainings were performed. The human MPM xenografts continue to develop on the CAMs and xenograft MPM cells showed highly metastatic features and a particular pattern of metastasis. The SaMNPs had a specific uptake by the MPM cells and an antiproliferative effect at therapeutic doses greater than 100 µg÷mL. The results confirmed the possibility to use the CAM as in vivo model to study the biology of MPM and to evaluate the antitumor potential of new therapeutic agents. They highlighted a strong antitumor effect of the SaMNPs on the rat and human MPM cells and open new perspectives in the treatment of MPM.

Simultaneous Quantitation of Boric Acid and Calcium Fructoborate in Dietary Supplements by HPTLC-Densitometry.

The paper describes a new, simple, selective and precise high-performance thin-layer chromatographic (HPTLC) method for the simultaneous identification and quantitative determination of boric acid (BA) and calcium fructoborate (CFB) in bulk and tablet/capsule dosage forms of dietary supplements. HPTLC silica gel G 60 F254 precoated glass plates were used as the stationary phase. The mobile phase consisted of 2-propanol-water 8:2 (v/v). The two boron-based compounds were adequately separated with the Rf values of 0.83 ± 0.01 (BA) and 0.59 ± 0.01 (CFB).

Calcium Fructoborate for Bone and Cardiovascular Health.

Calcium fructoborate (CF), a natural sugar-borate ester found in fresh fruits and vegetables, is a source of soluble boron. CF contains three forms of borate (diester, monoester, and boric acid) and all are biologically active, both at the intracellular (as free boric acid) and extracellular level (as fructose-borate diester and monoester). At the cellular and molecular level, CF is superior to the boric acid/borate, exhibiting a complex "protective" effect against inflammatory response. CF is commercially available in the USA as a "nature-identical" complex, an active compound for dietary supplements. It provides effective and safe support against the discomfort and lack of flexibility associated with osteoarticular conditions (arthritis and joint degeneration), and improves Western Ontario and McMaster Universities Osteoarthritis (WOMAC) and McGill indexes. In addition, orally administered CF is effective in ameliorating symptoms of physiological response to stress, including inflammation of the mucous membranes, discomfort associated with osteoarthritis disorders, and bone loss, and also for supporting cardiovascular health. Clinical studies have exhibited the ability of CF to significantly modulate molecular markers associated with inflammatory mechanisms, mainly on the elevated serum levels of C-reactive protein (CRP).

Development and Validation of an HPLC Method for Simultaneous Quantification of Clopidogrel Bisulfate, Its Carboxylic Acid Metabolite, and Atorvastatin in Human Plasma: Application to a Pharmacokinetic Study.

A simple, sensitive, and specific reversed phase liquid chromatographic method was developed and validated for simultaneous quantification of clopidogrel, its carboxylic acid metabolite, and atorvastatin in human serum. Plasma samples were deproteinized with acetonitrile and ibuprofen was chosen as internal standard. Chromatographic separation was performed on an BDS Hypersil C18 column (250 × 4.6 mm; 5 µm) via gradient elution with mobile phase consisting of 10 mM phosphoric acid (sodium) buffer solution (pH = 2.6 adjusted with 85% orthophosphoric acid) : acetonitrile : methanol with flow rate of 1 mL·min(-1). Detection was achieved with PDA detector at 220 nm. The method was validated in terms of linearity, sensitivity, precision, accuracy, limit of quantification, and stability tests. Calibration curves of the analytes were found to be linear in the range of 0.008-2 µg·mL(-1) for clopidogrel, 0.01-4 µg·mL(-1) for its carboxylic acid metabolite, and 0.005-2.5 µg·mL(-1) for atorvastatin. The results of accuracy (as recovery) with ibuprofen as internal standard were in the range of 96-98% for clopidogrel, 94-98% for its carboxylic acid metabolite, and 90-99% for atorvastatin, respectively.

Effects of Calcium Fructoborate on Levels of C-Reactive Protein, Total Cholesterol, Low-Density Lipoprotein, Triglycerides, IL-1ß, IL-6, and MCP-1: a Double-blind, Placebo-controlled Clinical Study.

Calcium fructoborate (CFB) has been reported as supporting healthy inflammatory response. In this study, we assess the effects of CFB on blood parameters and proinflammatory cytokines in healthy subjects. This was a randomized, double-blinded, placebo-controlled trial. Participants received placebo or CFB at a dose of 112 mg/day (CFB-1) or 56 mg/day (CFB-2) for 30 days. Glucose, total cholesterol (TC), low-density lipoprotein (LDL), high-density lipoprotein (HDL), triglycerides (TG), C-reactive protein (CRP), homocysteine, interleukin 1 beta (IL-1ß), IL-6, and monocyte chemoattractant protein-1 (MCP-1) were determined before and after supplementation. CFB-1 showed a reduction in blood levels of CRP by 31.3 % compared to baseline. CFB-1 and CFB-2 reduced LDL levels by 9.8 and 9.4 %, respectively. CFB-1 decreased blood homocysteine by 5.5 % compared with baseline, whereas CFB-2 did not have a significant effect. Blood levels of TG were reduced by 9.1 and 8.8 % for CFB-1 and CFB-2, respectively. Use of both CFB-1 and CFB-2 resulted in significantly reduced IL-6 levels, when compared within and between groups. IL-1ß was reduced by 29.2 % in the CFB-1 group. Finally, CFB-1 and CFB-2 reduced MCP-1 by 31 and 26 %, respectively. Our data indicate that 30-day supplementation with 112 mg/day CFB (CFB-1) resulted in a significant reduction of LDL, TG, TC, IL-1ß, IL-6, MCP-1, and CRP. HDL levels were increased, when compared to baseline and placebo. These results suggest that CFB might provide beneficial support to healthy cardiovascular systems by positively affecting these blood markers (ClinicalTrials.gov, ISRCTN90543844; May 24, 2012 ( http://www.controlled-trials.com/ISRCTN90543844 )).

Effect of propranolol on mepivacaine serum concentrations in dental practice.

OBJECTIVE: The objective of this study was to determine the effect of propranolol pretreatment on mepivacaine serum concentrations in dental patients. STUDY DESIGN: In a double blind, randomized, 2-way crossover study, 10 patients ingested 30 mg propranolol or placebo, 2 hours before local anesthesia for dental scaling. Each subject received a single dose of 51 mg mepivacaine for posterior superior alveolar nerve block. Mepivacaine in venous serum was measured for up to 1 hour, after 5, 15, 30, 45, and 60 minutes from injection. Serum concentrations of mepivacaine were determined by gas chromatography. Blood pressure and heart rate were measured before and after propranolol or placebo and after each sampling. RESULTS: Peak serum concentrations of mepivacaine, C(max) (1.214 +/- 0.746 microg/mL(-1)), were significantly increased by propranolol (2.249 +/- 1.559 microg/mL(-1), P < .05). Propranolol pretreatment reduced blood pressure and heart rate. CONCLUSION: Although propranolol pretreatment increased almost doublefold mepivacaine serum concentrations and reduced blood pressure and heart rate, mepivacaine can be used safely in dental patients taking propranolol for short-duration interventions.