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Željka Vanić,1 Zora Rukavina,1 Suvi Manner,2 Adyary Fallarero,3 Lidija Uzelac,4 Marijeta Kralj,4 Daniela Amidžić Klarić,1 Anita Bogdanov,5 Tímea Raffai,5 Dezső Peter Virok,5 Jelena Filipović-Grčić,1 Nataša Škalko-Basnet61Department of Pharmaceutical Technology, Faculty of Pharmacy and Biochemistry, University of Zagreb, 10000 Zagreb, Croatia; 2Pharmaceutical Sciences Laboratory, Faculty of Science and Engineering, Åbo Akademi and University of Helsinki, 20520 Turku, Finland; 3Division of Pharmaceutical Biosciences, Pharmaceutical Biology, Faculty of Pharmacy, University of Helsinki, 00014 Helsinki, Finland; 4Department of Molecular Medicine, Ruđer Bošković Institute, 10000 Zagreb, Croatia; 5Department of Medical Microbiology and Immunobiology, University of Szeged, 6720 Szeged, Hungary; 6Drug Transport and Delivery Research Group, Department of Pharmacy, Faculty of Health Sciences, University of Tromsø the Arctic University of Norway, 5037 Tromsø, NorwayBackground: Efficient localized cervicovaginal antibacterial therapy, enabling the delivery of antibiotic to the site of action at lower doses while escaping systemic drug effects and reducing the risk of developing microbial resistance, is attracting considerable attention. Liposomes have been shown to allow sustained drug release into vaginal mucosa and improve delivery of antibiotics to bacterial cells and biofilms. Azithromycin (AZI), a potent broad-spectrum macrolide antibiotic, has not yet been investigated for localized therapy of cervicovaginal infections, although it is administered orally for the treatment of sexually transmitted diseases. Encapsulation of AZI in liposomes could improve its solubility, antibacterial activity, and allow the prolonged drug release in the cervicovaginal tissue, while avoiding systemic side effects.Purpose: The objective of this study was to develop AZI-liposomes and explore their potentials for treating cervicovaginal infections.Methods: AZI-liposomes that differed in bilayer elasticity/rigidity and surface charge were prepared and evaluated under simulated cervicovaginal conditions to yield optimized liposomes, which were assessed for antibacterial activity against several planktonic and biofilm-forming Escherichia coli strains and intracellular Chlamydia trachomatis, ex vivo AZI vaginal deposition/penetration, and in vitro cytotoxicity toward cervical cells.Results: Negatively charged liposomes with rigid bilayers (CL-3), propylene glycol liposomes (PGL-2) and deformable propylene glycol liposomes (DPGL-2) were efficient against planktonic E. coli ATCC 700928 and K-12. CL-3 was superior for preventing the formation of E. coli ATCC 700928 and K-12 biofilms, with IC50 values (concentrations that inhibit biofilm viability by 50%) up to 8-fold lower than those of the control (free AZI). DPGL-2 was the most promising for eradication of already formed E. coli biofilms and for treating C. trachomatis infections. All AZI-liposomes were biocompatible with cervical cells and improved localization of the drug inside vaginal tissue compared with the control.Conclusion: The performed studies confirm the potentials of AZI-liposomes for localized cervicovaginal therapy.Keywords: vaginal drug delivery, biofilm, Escherichia coli, Chlamydia trachomatis, cervical cells, biocompatibility more...

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Background/objectives: Janus kinase inhibitors open new horizons for small-molecule drugs in treating inflammatory bowel disease, with ritlecitinib demonstrating significant efficacy in clinical trials for ulcerative colitis and Crohn's disease. Ritlecitinib, a second-generation JAK3 inhibitor, is a novel therapeutic agent for alopecia areata and other autoimmune conditions., Methods: A new stability-indicating UHPLC-DAD-MS/MS method was developed, validated, and applied for a forced degradation study of ritlecitinib under ICH guidelines., Results: The method demonstrated high specificity, sensitivity (LOD: 0.04 µg/mL; LOQ: 0.14 µg/mL), precision (RSD ≤ 0.15%), and accuracy (99.9-100.3%). Forced degradation studies under acidic, basic, oxidative, thermal, and photolytic conditions revealed four novel degradation products. Basic degradation followed second-order kinetics, while oxidative degradation followed zero-order kinetics., Conclusions: The validated method reliably characterized ritlecitinib's stability and degradation products, providing essential data for optimizing formulation, determining proper storage conditions, anticipating drug-excipient interactions, and ensuring quality control. The eco-friendliness and applicability of the developed forced degradation procedure were evaluated using various green and blue metric tools. Incorporating green analytical principles underscores its potential for sustainable pharmaceutical analysis. more...

Advanced oxidation processes (AOPs), including ionizing radiation treatment, are increasingly recognized as an effective method for the degradation of pharmaceutical pollutants, including non-steroidal anti-inflammatory drugs (NSAIDs). Nabumetone (NAB), a widely used NSAID prodrug, poses an environmental risk due to its persistence in aquatic ecosystems and its potential toxicity to non-target organisms. In this study, the radiolytic degradation of NAB was investigated under different experimental conditions (dose rate, radical scavenging, pH, matrix effect), and the toxicity of its degradation products was evaluated. NAB was rapidly degraded at 300 Gy with prolonged irradiation. Mineralization of about 88% of NAB solutions was observed based on the evaluation of total organic carbon (TOC). The most efficient degradation of NAB occurred under N 2 O conditions, while it was retarded in the presence of thiourea. The water matrix components had a significant influence on the efficiency of degradation. In addition, the main degradation products were identified by LC-HRMS. Toxicity studies on different bacteria showed no significant impact of the NAB degradation products, while in silico predictive methods revealed their slightly increased toxicity compared to the parent compound, but considerably lower toxicity in comparison to its main active form 6-methoxy-2-naphthylacetic acid (MNA). Additionally, significantly lower toxicities are predicted for degradation products in N 2 O saturated solution. These results underline the importance of optimizing irradiation parameters for effective degradation and minimizing the formation of harmful by-products. Understanding all aspects of the AOP processes and the toxicological effects of the degradation products ensures effective mitigation of potential environmental and health risks of water treatment processes., Competing Interests: The authors declare no conflicts of interest. more...

Background: Nabumetone (NAB) is a poorly soluble nonsteroidal anti-inflammatory prodrug (BCS class II drug) whose solubility is significantly improved by complexation with cyclodextrins (CDs). Methods : The solid complexes, in a 1:1 molar ratio, were prepared by mechanochemical activation by grinding, using β-cyclodextrin (β-CD) and its derivatives, hydroxypropyl- and sulfobutylether-β-cyclodextrin (HP-β-CD and SBE-β-CD). The complexation was confirmed by differential scanning calorimetry (DSC), powder X-ray diffraction (PXRD), and attenuated total reflectance Fourier-transformed infrared spectroscopy (ATR-FTIR). Obtained products were further characterized regarding their solubility, in vitro dissolution, permeability and chemical stability. Results : Co-grinding with HP-β-CD and SBE-β-CD yielded products that showed in vitro dissolution profiles in hydrochloric acid medium (pH 1.2) that were substantially different from that of pure NAB, yielding dissolution efficiency enhancements of 34.86 ± 1.64 and 58.30 ± 0.28 times, respectively, for the optimized products. Their in vitro dissolution and gastrointestinal permeability were also studied in a low-volume environment at pH 6.8, corresponding to the intestinal environment. Both β-CD derivatives increased NAB dissolution rate and NAB mass transport across the biomimetic membrane. The effect of β-CD derivatives on NAB chemical stability was studied under the stress conditions by the developed and validated UHPLC-DAD-HRMS method. In acidic conditions, pure and complexed NAB was prone to hydrolytic degradation, yielding one degradation product-pharmacologically inactive NAB metabolite. However, under the oxidative conditions at elevated temperatures, 10 NAB degradation products were identified from co-ground samples. All systems were stable during photo- and long-term stability studies. Conclusions : NAB complexes with HP-β-CD and SBE-β-CD are promising candidates for pharmaceutical product development. more...

Etrasimod, a novel selective sphingosine-1-phosphate receptor modulator, was recently approved by the U.S. Food and Drug Administration and the European Medicinal Agency for the treatment of moderately to severely active ulcerative colitis in adults. In this research, the forced degradation study as an integral part of drug product and packaging development, which generates data on degradation mechanisms, is published. The development and validation of the stability-indicating method using a superior high-performance liquid chromatography technique coupled with a diode array detector and tandem mass spectrometer was performed to support the forced degradation study and monitor the formation of degradation products. Etrasimod demonstrated good stability under elevated temperature and basic stress conditions, while acidic hydrolysis, oxidative, and photolytic degradation produced eight degradation products. The knowledge of degradation products will be useful in the long-term stability study for establishing the acceptance criteria of etrasimod as a drug substance and dosage form during quality control and stability assessment. The eco-friendliness of the developed forced degradation procedure was evaluated using various greenness appraisal tools. The green metric tools showed that the forced degradation procedure obeys eco-friendly conditions., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.) more...

We present a case of a healthy young male professional water polo player who presented with swelling and pain in the upper arm and elbow after vigorous exercise. Diagnostic workup included an MRI and dynamic duplex ultrasound, which revealed compression of the axillary vein by a hypertrophic pectoralis minor muscle without thrombosis, constituting McCleery syndrome. This is a rare entity within the multiple thoracic outlet syndrome aetiologies. Taking a detailed history and physical examination complemented with diagnostic imaging are vital to the diagnosis. Afterward, the patient was treated with multimodal physical therapy and fully recovered and even exceeded his previous training and play level. more...

This paper aims to evaluate the product contamination by elemental impurities during the mechanochemical synthesis of praziquantel (PZQ) co-crystal, polymeric dispersion and cyclodextrin complex by grinding. To assess that, PZQ was co-ground with malic acid (MA), Poloxamer F-127 (F-127) and hydroxypropyl-β-cyclodextrin (HPβCD) in high-energy vibrational mills using stainless steel and agate grinding tools, applying different processing time (30 and 90 min). Differential scanning calorimetry and X-ray powder diffraction confirmed the formation of the targeted products, regardless of applied processing time and grinding tool type. After digestion of the solid powder products, the levels of selected elemental impurities were analysed by inductively coupled plasma mass spectrometry (ICP-MS). The analysis revealed that the content of Mg, Ca, and V are below the limit of quantification in all samples analysed. The contents of P and Na are not related to the type of ball mill and reaction time, but to the starting materials themselves, considering that Na is found in HPβCD and MA, while P was found in F-127. The detected Si impurities in the co-ground products can be related to the use of the agate balls and jars, while the presence of Cr and Fe can be related to the use of the stainless steel grinding tools. The risk assessment showed that the oral administration of the prepared co-ground products in quantities corresponding to regular PZQ oral doses resulted in only insignificant exposure to Cr. Finally, the use of agate grinding tools should be preferred, as administration of such products results in lower Cr exposure. The presented elemental impurities did not lead to any significant drug degradation as PZQ content at the end of the six-month testing period was still in the range of 95-105 % of the initial content. Regardless, ICP-MS analysis of the elemental impurities should be considered in regular quality control procedures in the development and production of novel pharmaceutical products prepared by grinding., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.) more...

Glomerulonephritis following an enterococcal endocarditis is an extremely rare and life-threatening condition. We present the case of a 71-year-old patient with rapidly progressive glomerulonephritis following enterococcal endocarditis after surgical replacement of the aortic valve. The combination of antibiotic therapy, corticosteroid therapy and haemodialysis led to an improvement in renal function; however, the severity of cardiac deterioration resulted in a fatal outcome. more...

Inflammatory bowel disease is a complex disorder characterized by chronic gastrointestinal inflammation. Thus, patients prefer to use herbal dietary supplements containing turmeric, Indian frankincense, green chiretta, and black pepper in an attempt to cope better with their chronic condition. The dietary supplements' dosage forms and herbal ingredients were assessed in terms of the products' physicochemical parameters (weight uniformity, friability, disintegration, rupture test, tablet's breaking force, and powder flowability) in view of the USP-NF requirements. In addition, contaminants such as organic solvents and ethylene oxide were evaluated using gas chromatography. Assessment of gluten via an Enzyme-Linked Immunosorbent Assay was also performed. Most of the products met USP requirements. The high average weight of one multicomponent tablet sample with a high breaking force value can explain the observed negative results of the disintegration test. A total of 26% of samples tested positive for gluten, but the most alarming fact is that the ethylene oxide levels found in two samples were up to 30 times higher than the EU limit. Accordingly, dietary supplement quality control is of fundamental importance., Competing Interests: The authors declare no conflicts of interest. more...

Biocompatible mucoadhesive formulations that enable a sustained drug delivery at the site of action, while exhibiting inherent antimicrobial activity, are of great importance for improved local therapy of vaginal infections. The aim of this research was to prepare and evaluate the potential of the several types of azithromycin (AZM)-liposomes (180-250 nm) incorporated into chitosan hydrogel (AZM-liposomal hydrogels) for the treatment of aerobic vaginitis. AZM-liposomal hydrogels were characterized for in vitro release, and rheological, texture, and mucoadhesive properties under conditions simulating the vaginal site of application. The role of chitosan as a hydrogel-forming polymer with intrinsic antimicrobial properties was explored against several bacterial strains typical for aerobic vaginitis as well as its potential effect on the anti-staphylococcal activity of AZM-liposomes. Chitosan hydrogel prolonged the release of the liposomal drug and exhibited inherent antimicrobial activity. Additionally, it boosted the antibacterial effect of all tested AZM-liposomes. All AZM-liposomal hydrogels were biocompatible with the HeLa cells and demonstrated mechanical properties suitable for vaginal application, thus confirming their potential for enhanced local therapy of aerobic vaginitis. more...

Alport syndrome (AS) and thin basement membrane nephropathy (TBMN) are part of the spectrum of kidney disorders caused by pathogenic variants in α3, α4, or α5 chains of the collagen type IV, the major structural component of the glomerular basement membrane (GBM). Using targeted next-generation sequencing (NGS), 34 AS/TBMN patients (58.8% male) from 12 unrelated families were found positive for heterozygous c.2881+1G>A variant of the COL4A3gene, that is considered disease-causing. All patients were from the continental or island part of Croatia. Clinical, laboratory, and histopathological data collected from the medical records were analyzed and compared to understand the clinical course and prognosis of the affected patients. At the time of biopsy or first clinical evaluation, the mean age was 31 years (median: 35 years; range: 1 - 72 years). Hematuria was present in 33 patients (97.1%) and 19 (55.9%) patients had proteinuria. There were 6 (17.6%) patients with hearing loss, 4 (11.8%) with ocular lesions, and 11 (32.4%) with hypertension. Twenty-three (67.6%) patients had proteinuria at follow-up, and 5 (14.7%) patients with the median age of 48 years (range: 27-55) progressed to kidney failure, started dialysis, or underwent kidney transplantation. Of the 13 patients who underwent kidney biopsy, 4 (30.8%) developed focal segmental glomerulosclerosis (FSGS), and 8 (66.7%) showed lamellation of the GBM, including all patients with FSGS. It is essential to conduct a detailed analysis of each collagen type IV genetic variant to optimize the prognosis and therapeutic approach for affected patients. more...

Research Background: As use of functional food and herbal combination products is ever increasing, methods for quality control of such preparations are necessary. Moreover, low quality of products can cause either lack of benefit or harm to the consumer. In this work, determination of three curcuminoids, piperine, six boswellic acids and three andrographolides, often used in combination products, was carried out in raw materials and dietary supplements., Experimental Approach: After extraction optimization using Box-Behnken design, maximum active substance yields were obtained using 81.5% ethanol in hydroethanolic extraction solvent, 30 min sonication time and 60 °C extraction temperature. Afterwards, a high-performance liquid chromatography method was developed and validated, with special attention paid to selectivity, precision and robustness of the method. Lastly, 54 food and dietary supplement samples were analyzed., Results and Conclusions: Most products were bought locally, from credible vendors and they all complied with relevant regulatory requirements. However, products obtained on the Internet contained little to no active substances (24% of samples contained less than 20% declared content), presumably showing no efficacy, or were either found to be likely adulterated or contained very high amounts of active substances, compromising safety in terms of dose-dependent adverse effects (one sample containing andrographolides) and pharmacokinetic interactions (one sample containing piperine). In conclusion, consumers should refrain from purchasing such products from the Internet and obtain them only from verified suppliers such as local pharmacies or health stores., Novelty and Scientific Contribution: This work demonstrates the first developed method for the analysis of aforementioned combination products, which are on the rise today. The method is simple and robust and can be adapted by most laboratories for routine quality control of the said products. Moreover, the work sheds light on the low quality of several products and signifies the need for increased consumer awareness of dangers of taking such products., Competing Interests: CONFLICT OF INTEREST The authors declare no conflict of interest. more...

The in vitro antimicrobial activity of Fe(III) and Ga(III) complexes with N'-(2,3-dihydroxy-phenylmethylidene)-3-pyridinecarbohydrazide (H 2 L 1 ), N'-(2,4-dihydroxy-phenyl-methylidene)-3-pyridinecarbohydrazide (H 2 L 2 ), N'-(2,5-dihydroxy-phenylmethylidene)-3-pyridinecarbohydrazide (H 2 L 3 ), N'-(2-hydroxy-3-methoxyphenyl-methylidene)-3-pyridine-carbohydrazide (H 2 L 4 ), N'-(2-hydroxy-4-methoxyphenylmethyl-idene)-3-pyridine-carbohydrazide (H 2 L 5 ), and N'-(2-hydroxy-5-methoxyphenylmethylidene)-3-pyridinecarbo-hydrazide (H 2 L 6 ) toward several Gram-positive strains of Staphylococcus aureus, a Gram-negative strain of Escherichia coli, and a yeast Candida albicans were investigated. Fe(III)-complexes do not possess antimicrobial activity against all tested strains at concentrations up to 10 mg mL -1 . Ga(III) complexes with dihydroxy derivatives showed selective activity, while the broadest range of antibacterial and antifungal activities was observed for complex with 2-hydroxy-3-methoxy-derivative, ligand H 2 L 5 . In addition, the coordination properties of ligands H 2 L 1 -H 2 L 3 in solution were investigated by UV-Vis spectroscopy. The stability constants (logK) for Ga(III)-H 2 L 1:1 complexes in MeOH/H 2 O 1/1 at pH 2.52 were determined, and amounted to 5.8, 5.68, and 4.7, respectively. Detailed characterization of complexes was performed by high-resolution mass spectrometry. The fragmentation pathways for dimer [Fe 2 (L 1 ) 2 ] 2+ , [Fe(HL) 2 ] + , [Ga(HL 2 ) 2 ] + and adduct ions are given. The comparison with analogue Ga(III) and Fe(III) complexes with compounds H 2 L 4 -H 2 L 6 was made as well., (© 2022. The Author(s), under exclusive licence to Society for Biological Inorganic Chemistry (SBIC).) more...

Collapsing glomerulopathy (CG) or collapsing focal segmental glomerulosclerosis (cFSGS) is an aggressive disease with a high tendency of progression to end-stage renal disease due to common resistance to conventional immunosuppressants. Rituximab (RTX), a monoclonal antibody against CD20 B cells, showed some benefit in the treatment of CG. We are reporting about female patients with an idiopathic form of CG presenting with nephrotic syndrome (NS) and renal insufficiency resistant to several immunosuppressive agents such as steroids (ST), calcineurin inhibitors (CNI), and cyclophosphamide (CYC). This multidrug-resistant disease responded to RTX with complete remission. Forty-four months after initial RTX administration, a relapse of CG with severe NS and acute renal insufficiency occurred. Repeated application of RTX led to complete remission again. To the best of our knowledge, we are reporting the first case of the relapsing multidrug-resistant form of CG, which responded to RTX. Current data about the treatment of CG with RTX is lacking and is based on rare case reports and small case series. Thus, our report can contribute to determining the role of RTX in the treatment of CG. more...

In the present study, various procedures have been compared for the determination of lipophilicity, hydrophobicity, and plasma protein binding of curcuminoids, boswellic acids, andrographolides, and piperine as biologically active ingredients of botanicals used in IBD treatment. Our results have shown that IAM-HPLC assay is the most suitable one for lipophilicity determination of all analytes regardless of their class and botanical source. HSA-HPAC and AGP-HPAC assays revealed that all investigated compounds have a higher affinity for HSA which is the most abundant protein in human plasma. The high affinity of biologically active compounds to all biological structures (phospholipids and proteins) admonishes that their small portion is available for therapeutic effects in IBD patients. Our experimental research is complemented by various theoretical approaches based on different algorithms for pharmacokinetic properties prediction. The similarities between experimental and calculated values were evaluated using PCA and CA as a statistical tool. The statistical analysis implies that plasma protein binding is a complex process, and theoretical approaches still cannot fully replace experimental ones. more...

Since oxidative stress has been linked to several pathological conditions and diseases, drugs with additional antioxidant activity can be beneficial in the treatment of these diseases. Therefore, this study takes a new look at the antioxidant activity of frequently prescribed drugs using the HPLC-DPPH method. The antioxidative activity expressed as the TEAC value of 82 drugs was successfully determined and is discussed in this work. Using the obtained values, the QSAR model was developed to predict the TEAC based on the selected molecular descriptors. The results of QSAR modeling showed that four- and seven-variable models had the best potential for TEAC prediction. Looking at the statistical parameters of each model, the four-variable model was superior to seven-variable. The final model showed good predicting power ( r = 0.927) considering the selected descriptors, implying that it can be used as a fast and economically acceptable evaluation of antioxidative activity. The advantage of such model is its ability to predict the antioxidative activity of a drug regardless of its structural diversity or therapeutic classification. more...

In this work we present the development of in situ gelling nanosuspension as advanced form for fluticasone propionate nasal delivery. Drug nanocrystals were prepared by wet milling technique. Incorporation of drug nanocrystals into polymeric in situ gelling system with pectin and sodium hyaluronate as constitutive polymers was fine-tuned attaining appropriate formulation surface tension, viscosity and gelling ability. Drug nanonisation improved the release profile and enhanced formulation mucoadhesive properties. QbD approach combining formulation and administration parameters resulted in optimised nasal deposition profile, with 51.8% of the dose deposited in the middle meatus, the critical region in the treatment of rhinosinusitis and nasal polyposis. Results obtained in biocompatibility and physico-chemical stability studies confirmed the leading formulation potential for safe and efficient nasal corticosteroid delivery., (Copyright © 2022 Elsevier B.V. All rights reserved.) more...

Praziquantel (PZQ) is a biopharmaceutical classification system (BCS) class II anthelmintic drug characterized by poor solubility and a bitter taste, both of which can be addressed by inclusion complexation with cyclodextrins (CD). In this work, a comprehensive investigation of praziquantel/cyclodextrin (PZQ/CD) complexes was conducted by means of UV-vis spectroscopy, spectrofluorimetry, NMR spectroscopy, liquid chromatography-high-resolution mass spectrometry (LC-HRMS/MS), and molecular modeling. Phase solubility studies revealed that among four CDs tested, the randomly methylated β-CD (RMβCD) and the sulfobutylether sodium salt β-CD (SBEβCD) resulted in the highest increase in PZQ solubility (approximately 16-fold). The formation of 1:1 inclusion complexes was confirmed by HRMS, NMR, and molecular modeling. Both cyclohexane and the central pyrazino ring, as well as an aromatic part of PZQ are included in the CD central cavity through several different binding modes, which exist simultaneously. Furthermore, the influence of CDs on PZQ stability was investigated in solution (HCl, NaOH, H 2 O 2 ) and in the solid state (accelerated degradation, photostability) by ultra-high-performance liquid chromatography-diode array detection-tandem mass spectrometry (UPLC-DAD/MS). CD complexation promoted new degradation pathways of the drug. In addition to three already known PZQ degradants, seven new degradation products were identified ( m / z 148, 215, 217, 301, 327, 343, and 378) and their structures were proposed based on HRMS/MS data. Solid complexes were prepared by mechanochemical activation, a solvent-free and ecologically acceptable method. more...

Nasal route of administration offers a unique opportunity of brain targeted drug delivery via olfactory and trigeminal pathway, providing effective CNS concentrations at lower doses and lower risk for adverse reactions compared to systemic drug administration. Therefore, it has been recently proposed as a route of choice for glucocorticoids to control neuroinflammation processes in patients with severe Covid-19. However, appropriate delivery systems tailored to enhance their efficacy yet need to emerge. In this work we present the development of sprayable brain targeting powder delivery platform of dexamethasone sodium phosphate (DSP). DSP-loaded microspheres, optimised employing Quality-by-Design approach, were blended with soluble inert carriers (mannitol or lactose monohydrate). Powder blends were characterized in terms of homogeneity, flow properties, sprayability, in vitro biocompatibility, permeability and mucoadhesion. Nasal deposition studies were performed using 3D printed nasal cavity model. Mannitol provided better powder blend flow properties compared to lactose. Microspheres blended with mannitol retained or enlarged their mucoadhesive properties and enhanced DSP permeability across epithelial model barrier. DSP dose fraction deposited in the olfactory region reached 17.0% revealing the potential of developed powder platform for targeted olfactory delivery. The observed impact of nasal cavity asymmetry highlighted the importance of individual approach when aiming olfactory region. more...

Adherence in chronic diseases is a major problem which can be combated by prescribing fixed-dose combinations in the therapy of the disease. Thus, a combination of azathioprine and folic acid in the treatment of inflammatory bowel disease is highly required, but prior to formulation development, chemical compatibility of the two drugs needs to be investigated. In this work, differential scanning calorimetry, isothermal stress testing, in vitro dissolution and forced degradation studies were utilized to investigate compatibility. Moreover, a stability-indicating HPLC-DAD method for the determination of parent drugs and five of their impurities was developed, validated and applied to the in-house sample. Compatibility testing revealed no noteworthy interactions of the two drug substances. Furthermore, forced degradation showed no substantial differences between the degradation profiles of each active pharmaceutical ingredient, their mixture and the in-house sample, further reinforcing the claim of compatibility. Lastly, the in-house sample was analyzed: it was shown to conform to the requirements of relevant regulatory documents for all the investigated analytes, demonstrating the method's viability for use in formulation and process development. Our results give way to the possibility of realization of said fixed-dose combination. more...

The simultaneous administration of sulfasalazine and folic acid is regular practice in the therapy of inflammatory bowel diseases in order to maintain sufficient folate concentration in patients. Having multiple drugs in the therapy increases the possibility of patients failing adherence, thus unintentionally endangering their health. A fixed-dose combination of sulfasalazine and folic would simplify the classical polytherapeutic approach; however, the physicochemical compatibility investigation of two active pharmaceutical ingredients plays an important role in the development of such a product. In this work, various analytical tools were used to determine the physicochemical compatibility of sulfasalazine and folic acid. For the evaluation of chemical compatibility, infrared spectroscopy in combination with advanced statistical methods, such as the principal component analysis and cluster analysis, were used, whilst a simultaneous thermogravimetry/differential thermal analysis gave us an insight into the physical compatibility of two drugs. Isothermal stress testing, forced degradation and dissolution studies, followed by the analysis with a developed chromatographic method for the monitoring of folic acid, sulfasalazine and two of its related impurities, sulfapyridine and salicylic acid, gave us an insight into its chemical compatibility. The combination of the results obtained from the used techniques implies a satisfactory physicochemical compatibility between sulfasalazine and folic acid, which opens the path to the development of the proposed fixed-dose combination. more...

In this work, a systematical compatibility investigation of 6-mercaptopurine and folic acid, two commonly used medications in the treatment of inflammatory bowel disease, for the needs of a fixed-dose combination development strategy is shown. Various techniques and approaches, such as differential scanning calorimetry, isothermal stress testing, attenuated total reflectance-Fourier-transform infrared spectroscopy, dissolution medium stability and forced degradation studies, were used to elucidate the possible interactions from different aspects. The results predominantly point to the absence of physicochemical interactions between the examined substances in a variety of possible conditions. However, the forced degradation of the blend of substances and excipients in basic conditions showed a drastic degradation of 6-mercaptopurine, signifying that attention needs to be directed to the careful selection of the excipients for the formulation. To sum up, our findings indicate that a fixed-dose combination of 6-mercaptopurine and folic acid could be produced using one formulation blend, immensely simplifying its manufacture. more...

Inflammatory bowel disease is a common name for Crohn's disease and ulcerative colitis. These inflammatory states cause damage in the sidewalls of the gastrointestinal tract, resulting in malabsorption of food and vitamins. Folic acid (Vitamin B9) is often associated with inflammatory bowel diseases since reduced overall folate concentration in the human body may lead to the development of colorectal cancer and megaloblastic anaemia. However, its deficiency is easily compensated by taking an additional folic acid pill during regular therapy. At the moment, there are no studies that have examined the compatibility of folic acid with 5-aminosalicylate drugs used in the treatment of inflammatory bowel diseases. In this work, differential scanning calorimetry, forced degradation studies, isothermal stress testing and dissolution stability testing were used to determine the stability of folic acid and one of the most commonly used 5-aminosalicylates, mesalazine, when present in the same solution or blend. To monitor the assay of folic acid, mesalazine and nine of its related impurities, a single HPLC method was developed. Results of compatibility studies showed that no physicochemical interaction between mesalazine and folic acid occurs when combined, opening the path to the development of new formulations, such as a mesalazine/folic acid fixed-dose combination. more...

With the increase in the number of medicines patients have to take, there has been a rapid rise of fixed-dose combinations (FDCs) in the last two decades. Prior to FDC development, pharmacokinetic properties of active pharmaceutical ingredients (APIs) have to be evaluated, as well as methods for their determination developed. So as to increase patient compliance in inflammatory bowel disease, three novel FDCs of thiopurine immunosuppressants and folic acid are proposed; physico-chemical and pharmacokinetic properties such as hydrophobicity, lipophilicity and plasma protein binding of all APIs are evaluated. Moreover, experimental results of different properties are compared to those computed by various on-line prediction platforms so as to evaluate the viability of the in silico approach. A simultaneous method for their determination is developed, optimized, validated and applied to commercial tablet formulations. The method has shown to be fast, selective, accurate and precise, showing potential for reliable determination of API content in proposed FDCs during its development., Competing Interests: The authors declare no conflict of interest. more...

The aim of this study was to evaluate the implementation of the 9th edition of the American College of Chest Physicians (ACCP9) guidelines for prevention of venous thromboembolism in nonsurgical patients in clinical practice in one university and one general Croatian hospital. A retrospective study was conducted at Zadar General Hospital from Zadar and Dubrava University Hospital from Zagreb. Medical charts of all patients admitted to Medical Departments in two periods, before and after implementation of the ACCP9 guidelines, were analyzed. The ACCP9 guidelines were made available to all physicians through the hospital electronic information system immediately after the publication. The Hospital Drug Committees promoted implementation of the guidelines during their periodical clinical visits. Overall, 850 patients were included in the study in two periods. There was no statistically significant difference in the number of high-risk patients receiving thromboprophylaxis after the guidelines implementation in either hospital. In both periods, a signifi-cantly higher number of high-risk patients received thromboprophylaxis in Dubrava University Hos-pital in comparison with Zadar General Hospital (31.7% vs. 3.8% and 40.3% vs. 7.3%, respectively; p<0.001). This study revealed insufficient implementation of evidence-based thromboprophylaxis guidelines in clinical practice in two Croatian hospitals. more...

Malnutrition, inflammation, and anemia are common in peritoneal dialysis (PD) patients. In this study, correlations between Malnutrition Inflammation Score (MIS), laboratory and anthropometric parameters, and anemia indices in Croatian PD patients were analyzed. One hundred and one PD patients (males/females 54/47, age 58.71 ± 14.68 years, mean PD duration 21.82 ± 21.71 months) were included. Clinical, laboratory, and anthropometric parameters were measured. Statistically significant correlations between MIS and erythropoietin weekly dose per kg of body weight (ESA weekly dose), hemoglobin (Hb), and erythrocytes were found ( r = 0.439, p < 0.001; r = -0.032, p < 0.001; r = -0.435, p < 0.001), respectively. Also, statistically significant correlations were found between MIS and mean corpuscular volume ( r = 0.344, p < 0.001), iron ( r = -0.229, p = 0.021), and total iron binding capacity (TIBC) ( r = -0.362, p < 0.001), respectively. Furthermore, statistically significant correlations between ESA weekly dose and serum albumin level and body mass index (BMI) were found ( r = -0.272, p = 0.006; r = -0.269, p = 0.006), respectively. When we divided PD patients into 2 groups according Hb level (Hb ≥ 110 [ N = 60, 59.41 %]) and Hb < 110 [ N = 41, 40.59%]), statistically significant differences were found in MIS score (3.02 ± 2.54 vs 4.54 ± 3.54, p = 0.014), C-reactive protein (CRP) (3.52 ± 6.36 vs 7.85 ± 7.96, p = 0.005), and serum albumin level (44.22 ± 8.54 vs 39.94 ± 8.56, p = 0.003), respectively. Our findings suggest that anemia is correlated with malnutrition and inflammation in Croatian PD patients. Further studies are needed to assess whether modulating inflammatory or nutritional processes can improve anemia management in PD patients., (Copyright © 2017 International Society for Peritoneal Dialysis.) more...

Purpose: Levothyroxine (LT4) is a drug with a narrow therapeutic index, applied in small amounts (micrograms), which makes interactions in the absorption phase clinically significant. The main aim of this article was to review and present the latest information on factors that affect the gastrointestinal absorption of this drug., Methods: Relevant data were collected by using the MEDLINE, PubMed, EMBASE, Web of Science, Science Direct, and Scopus databases with the key words levothyroxine and absorption. Searches were not limited to specific publication types, study designs, dates, or languages. The reports were highly variable in the amount of information provided regarding study design and methods. Because of the heterogeneity of studies, no statistical analysis was performed., Findings: Many gastrointestinal disorders, such as celiac disease, atrophic gastritis, lactose intolerance, and Helicobacter pylori infection, may impede the absorption of levothyroxine. During treatment of these disorders, it is necessary to monitor serum thyroid-stimulating hormone and free T4 values to reduce the risk of developing iatrogenic hyperthyroidism. Soybeans and coffee have the greatest impact on the reduction of absorption, whereas vitamin C has the ability to increase it. Conversely, the effect of dietary fiber on the absorption of LT4 is not yet fully understood; further research is needed on this topic. A decrease in the absorption of LT4 is established and clinically significant when administered concomitantly with cholestyramine, colesevelam, lanthanum, calcium carbonate, calcium citrate, calcium acetate, iron sulfate, ciprofloxacin, aluminum hydroxide, sevelamer, or proton pump inhibitors. This effect should be taken into consideration when prescribing these drugs concomitantly with LT4. The effects of Giardia lamblia infection and the influence of orlistat, polystyrene sulfonate, raloxifene, and simethicone on absorption of LT4 have been poorly documented. For bariatric surgery, sucralfate and H 2 -antagonist interactions are not well founded or contradictory evidence is available regarding their existence; additional research should be conducted., Implications: The majority of the interactions are clinically significant. They are based on the LT4 adsorption on interfering substances in the digestive tract, as well as a consequently reduced amount of the drug available for absorption. These interactions can be avoided by separating the administration of LT4 and the interfering substance., (Copyright © 2017 Elsevier HS Journals, Inc. All rights reserved.) more...

Aims: This study was aimed at comparing the incidence of arterial hypertension and blood pressure (BP) variance in hospital and out-of-hospital hemodialysis (HD) patients during HD sessions., Methods: A cross-sectional study was conducted for 1 week at all the HD centers in Dalmatia, Croatia. The pre-, intra-, and post-dialysis BP values were collected for 3 consecutive HD sessions per patient., Results: Of the 399 subjects, 73.9% were hypertensives, who showed higher interdialytic weight gain compared to the normotensives (2.58 vs. 2.40). Hospital and out-of-hospital HD patients received identical antihypertensive therapies, except that beta blockers were more frequently administered to out-of-hospital HD patients. Higher pre-, intra-, and post-dialysis BP values were recorded in patients at out-of-hospital HD centers., Conclusion: The differences in BP variability and antihypertensive therapies administered to hospital HD patients as compared to out-of-hospital HD patients may reflect differing approaches by the nephrologists at these centers., (© 2017 S. Karger AG, Basel.) more...

Chronic kidney disease is clearly defined as a state of damaged kidney function lasting for more than three months. Changes manifest in serum and urine pathological findings with frequent morphological changes in the kidneys and reduction in glomerular filtration. The aim is to show the possibilities of renal replacement therapy and waste related disease during dialysis treatment. The methods are based on strong evidence and guidelines. Glomerular filtration is the basis in evaluating the stage of chronic kidney disease. Based on the measures of glomerular filtration reduction, chronic kidney disease is classified into five stages, thus facilitating approach to treatment of particular groups of patients depending on the level of glomerular filtration damage. Kidney function can be replaced by dialysis or transplantation and in certain cases symptomatically if the patient refuses dialysis treatment. Malnutrition, hypertension, kidney anemia and bone-mineral disease are often present in patients with higher stages of chronic kidney disease, particularly stage 5 and kidney function replacement by dialysis. In conclusion, timely treatment reduces morbidity and mortality in patients with chronic kidney disease. more...

Cardiorenal syndrome, a complex pathophysiological disorder of both the heart and kidneys, is a condition in which acute or chronic damage to one organ can lead to acute or chronic dysfunction of the other organ. Depending on primary organ dysfunction and disease duration, there are five different types of cardiorenal syndrome. Type 1 cardiorenal syndrome (acute cardiorenal syndrome) is defined as acute kidney injury caused by sudden decrease in heart function. Type 2 cardiorenal syndrome (chronic cardiorenal syndrome) refers to chronic kidney disease linked to chronic heart failure. Type 3 cardiorenal syndrome (acute renocardial syndrome) is caused by acute kidney injury that leads to heart failure. Type 4 cardiorenal syndrome (chronic renocardial syndrome) includes chronic heart failure due to chronic kidney disease. Type 5 cardiorenal syndrome (secondary cardiorenal syndrome) is reversible or irreversible condition marked by simultaneous heart and kidney insufficiency, as a result of multiorgan disease such as sepsis, diabetes mellitus, sarcoidosis, amyloidosis, etc. The pathophysiological patterns of cardiorenal syndrome are extremely complicated. Despite numerous publications, perplexed physiological, biochemical and hormonal disturbances as parts of the main pathogenic mechanisms of cardiorenal syndrome remain obscure. Even though there are guidelines for the treatment of patients with heart failure and chronic kidney disease, similar guidelines for the treatment of cardiorenal syndrome are lacking. In everyday practice, it is crucial to diagnose cardiorenal syndrome and use all diagnostic and therapeutic procedures available to prevent or alleviate kidney and heart failure. more...

Despite efforts by many dietary supplements' manufactures to reduce or replace ethanol, many products containing ethanol in concentrations up to 70% are available on market. Furthermore, botanical dietary supplements can vary in metal content as a function of the environment, processing equipment and product containers. Therefore, the aim of study was to develop a new rapid and highly sensitive method for simultaneous determination of ethanol and its impurities in dietary supplements by sHSS-GC-FID technique. In addition, contamination with metals by GFAAS technique was evaluated. The proposed sHSS-GC-FID method was successfully applied for analysis of 93 samples containing various amounts of ethanol. It should be highlighted that the dramatic variation from manufacture's claims was found in even one third of products. Furthermore, high amounts of ethanol were found in several products especially designed for children and in one product labeled as "alcohol-free". Metal impurities were below the limits established by USP., (Copyright © 2016 Elsevier Ltd. All rights reserved.) more...

Chronic kidney disease (CKD) is a systemic disease with numerous complications associated with increased morbidity and mortality. Chronic kidney disease-metabolic bone disease (CKD-MBD) starts at early stages of CKD with phosphorus accumulation and consequent initiation of numerous events that result with the development of secondary hyperparathyroidism with changes on bones and extraskeletal tissues. The most important and clinically most relevant consequences of CKD-MBD are vascular calcifications which contribute to cardiovascular mortality. Patients with the increased risk for the development of CKD-MBD should be recognized and treated. Prevention is the most important therapeutic option. The first step should be nutritional counseling with vitamin supplementation if necessary and correction of mineral status. Progression of CKD requires more intensive medicamentous treatment with the additional correction of metabolic acidosis and anemia. Renal replacement therapy should be timely initiated, with the adequate dose of dislaysis. Ideally, preemptive renal transplantion should be offered in individuals without contraindication for immunosuppressive therapy. more...

This study brings out the data on the content of 21 mineral and heavy metal in 15 blackberry wines made of conventionally and organically grown blackberries. The objective of this study was to classify the blackberry wine samples based on their mineral composition and the applied cultivation method of the starting raw material by using chemometric analysis. The metal content of Croatian blackberry wine samples was determined by AAS after dry ashing. The comparison between an organic and conventional group of investigated blackberry wines showed statistically significant difference in concentrations of Si and Li, where the organic group contained higher concentrations of these compounds. According to multivariate data analysis, the model based on the original metal content data set finally included seven original variables (K, Fe, Mn, Cu, Ba, Cd and Cr) and gave a satisfactory separation of two applied cultivation methods of the starting raw material. more...

There is a high incidence of cardiovascular morbidity and mortality among patients with chronic kidney disease (CKD) and malnutrition is a powerful predictor of cardiovascular morbidity and mortality in this population of patients. A multitude of factors related to CKD and renal replacement therapy can affect the nutritional status of CKD patients and lead to the development of malnutrition. In patients with CKD, protein energy wasting (PEW) is a condition that is distinct from undernutrition and is associated with inflammation, increased resting energy expenditure, low serum levels of albumin and prealbumin, sarcopenia, weight loss and poor clinical outcomes. Nutritional and metabolic derangements are implicated for the development of PEW in CKD and leading to the development of chronic catabolic state with muscle and fat loss. Prevention is the best way in treating PEW. Appropriate management of CKD patients at risk for PEW requires a comprehensive combination of strategies to diminish protein and energy depletion, and to institute therapies that will avoid further losses. The mainstay of nutritional treatment in MHD patients is nutritional counselling and provision of an adequate amount of protein and energy, using oral supplementation as needed. Intradialytic parenteral nutrition and total enteral nutrition should be attempted in CKD patients who cannot use the gastrointestinal tract efficiently. Other strategies such as anemia correction, treatment of secondary hyperparathyroidism and acidosis, delivering adequate dialysis dose can be considered as complementary therapies in CKD patients. Multidisciplinary work of nephrologists, gastroenterologist and dietician is needed to achieve best therapeutic goals in treating CKD patients with PEW. more...

In our clinical practice, we are often faced with emotional difficulties of transplanted patients. Most are due to anxiety, difficulty in integrating newly recruited organ as its own, feeling of guilt, and difficulties with personal experience of self. Despite common presence of emotional difficulties, many studies describe improvement in the quality of life of transplant patients. However, the quality of life is deteriorating again in case of transplant rejection. In such situations, restlessness develops along with losing control over their own lives, a sense of failure, hopelessness and lack of prosperity ideas. Complex emotional experiencing is very important in strengthening the patient's psychological health and personality, thus achieving better treatment compliance in general. more...

According to the National Registry of Renal Replacement Therapy (RRT), the incidence of chronic kidney disease (end-stage renal disease) and the need of RRT have declined in the last decade renal. One of the reasons for this tendency certainly is transplantation as the best choice. However, transplant procedure has limitations in elderly patients due to the number of comorbidities. This study was designed as retrospective analysis of outcomes in patients treated with peritoneal dialysis for a period of eleven years. Patients were divided into those who had been assisted or unassisted. Out of 100 patients treated with peritoneal dialysis (PD), 77 completed the treatment, including 26 assisted and 51 unassisted patients. Peritonitis was recorded in 20 assisted and 26 unassisted patients. Peritonitis was more common in unassisted patients, who were more frequently lost from PD. Assisted PD could be a good and safe choice of RRT in this special group of patients. more...