Levobunolol-imprinted polymer: a theoretical study.

CONTEXT: Levobunolol is a ß-blocker drug prescribed for the control and prevention of cardiovascular events, such as individuals with cardiac arrhythmia or a history of myocardial infarction. Creating a levobunolol-specific molecularly imprinted polymer (MIP) allows for enhanced selectivity, efficient sample preparation, controlled drug delivery, and improved sensing and detection capabilities. In this sense, the aim of this study was to obtain through DFT calculations the synthesis protocol of a MIP for levobunolol testing different functional monomers (FMs), solvents, and cross-linker agents (CLAs). The analysis of structural and energetic data led to the identification of the optimal MIP synthesis parameters, which involves the use of (trifluoromethyl)-arylic acid (TFMAA) as the functional monomer, toluene and chloroform as the solvents, and pentaerythritol triacrylate (PETRA) as the cross-linking agent. This rational design offers valuable insights for experimentalists seeking to efficiently synthesize a MIP for this important ß-blocker drug. METHODS: DFT calculations were conducted using the B97D functional along with the Pople's split valence 6-31G(d,p) basis set, which includes polarization functions on all atoms (B97D/6-31G(d,p)).

Identification of potential biomarkers and pathogenesis in neutrophil-predominant severe asthma: A comprehensive bioinformatics analysis.

BACKGROUND: Airway neutrophilia has been associated with asthma severity and asthma exacerbations. This study attempted to identify biomarkers, pathogenesis, and therapeutic molecular targets for severe asthma in neutrophils using bioinformatics analysis. METHODS: Fifteen healthy controls and 3 patients with neutrophilic severe asthma were screened from the Gene Expression Omnibus (GEO) database. Based on the analysis of differentially expressed genes (DEGs), functional and pathway enrichment analyses, gene set enrichment analysis, protein-protein interaction network construction, and analysis were performed. Moreover, small-molecule drug candidates have also been identified. RESULTS: Three hundred and three upregulated and 59 downregulated genes were identified. Gene ontology function enrichment analyses were primarily related to inflammatory response, immune response, leukocyte migration, neutrophil chemotaxis, mitogen-activated protein kinase cascade, Jun N-terminal kinase cascade, I-kappaB kinase/nuclear factor-κB, and MyD88-dependent toll-like receptor signaling pathway. Pathway enrichment analyses and gene set enrichment analysis were mainly involved in cytokine-cytokine receptor interaction, the TNF signaling pathway, leukocyte transendothelial migration, and the NOD-like receptor signaling pathway. Furthermore, 1 important module and 10 hub genes (CXCL8, TLR2, CXCL1, ICAM1, CXCR4, FPR2, SELL, PTEN, TREM1, and LEP) were identified in the protein-protein interaction network. Moreover, indoprofen, mimosine, STOCK1N-35874, trapidil, iloprost, aminoglutethimide, ajmaline, levobunolol, ethionamide, cefaclor, dimenhydrinate, and bethanechol are potential drugs for the treatment of neutrophil-predominant severe asthma. CONCLUSION: This study identified potential biomarkers, pathogenesis, and therapeutic molecular targets for neutrophil-predominant severe asthma.

Superior oblique myokymia treated with levobunolol.

Superior oblique myokymia (SOM) is an uncommon condition of unclear etiology that results in episodes of oscillopsia and diplopia. There is no established treatment protocol for SOM. We present 2 cases of SOM successfully managed with topical levobunolol 0.5%; both patients responded to a short course of medication administration and required minimal ongoing therapy. Case 1 was a 69-year-old woman with left SOM who had previously undergone a left Harada-Ito procedure. Her SOM improved immediately on administration of levobunolol and was maintained at follow-up 1 year later. Case 2 was a 49-year-old man with right SOM that affected his ability to work. After 2 days of topical levobunolol 0.5% nightly in the right eye, SOM episodes ceased; he continues to use drops intermittently for occasional recurrences.

Validated HPLC-DAD Method for the Simultaneous Determination of Six Selected Drugs Used in the Treatment of Glaucoma.

This work presents a simple, sensitive, and generic HPLC-diode-array detection method for the simultaneous determination of six drugs prescribed for the treatment of open-angle glaucoma and ocular hypertension. The investigated drugs include brimonidine tartarate (BMN), acetazolamide (AZA), brinzomaide (BZA), dorzolamide HCl (DZA), levobunolol HCl (LVB), and timolol maleate (TIM). Efficient chromatographic separation was achieved using a Thermo Hypersil BDS C18 column (4.6 × 250 mm, 5 µm) with a mobile phase consisting of phosphate buffer pH 5 and acetonitrile in a ratio of 78 + 22. The flow rate was 1 mL/min, and quantification was based on measuring peak areas at 298 nm for TIM and 254 nm for the other drugs. Peaks were perfectly resolved, with retention times at 3.06, 3.87, 4.53, 5.78, 7.31, and 10.78 min for BMN, AZA, DZA, TIM, LVB, and BZA respectively. The developed method was validated according to International Conference on Harmonization guidelines with respect to system suitability, linearity, ranges, accuracy, precision, robustness, and LODs and LOQs. The proposed method showed good linearity in the ranges of 2-80, 2.5-100, 2.5-100, 5-200, 3.75-150, and 1.75-70 µg/mL for BMN, AZA, DZA, TIM, LVB, and BZA, respectively. LODs were 0.20-1.01 µg/mL for the analyzed compounds. Applicability of the proposed method to real-life situations was assessed through the analysis of five different pharmaceutical formulations, and satisfactory results were obtained.

Effects of timolol maleate, levobunolol and apraclonidine on intraocular pressure, pupil size, blood pressure and heart rate in beagles / Efeitos do maleato de timolol, do levobunolol e da apraclonidina sobre a pressão intraocular, o diâmetro pupilar, a pressão sanguínea e a frequência cardíaca, em cães da raça Beagle

The aim of this study was to evaluate changes in intraocular pressure (IOP), pupil size (PS), blood pressure (BP), heart rate (HR), and ECG variables (Pms wave PmV, PR interval, QRS complex, RMV wave and QT intervals) over time during the instillation of 0.5% timolol, 0.5% levobunolol and 0.5% apraclonidine in clinically normal dogs. Ten adult beagles were used. Baseline values were measured at 8a.m., 2p.m. and 8p.m., for three consecutive days. A waiting period of 10 days between the administrations of each drug was established. For 15 consecutive days, the drug being tested was instilled in one eye of each dog twice a day (7a.m. and 7p.m.). The parameters were evaluated at the aforementioned times on days 3, 6, 9, 12 and 15. Data were statistically compared using the Bonferroni test and one-way repeated measures analysis of variance (P<0.05). The Pearson test was used to evaluate any correlation between QT interval, HR and BP. The tested drugs did not find a decrease in IOP. A significant decreased in PS was observed in almost all dogs following levobunolol administration, relative to the control eye. A significant decrease in HR was observed on day 3 following levobunolol treatment, while apraclonidine induced an increase on day 15. Blood pressure was reduced in all measurement time points following apraclonidine treatment. A negative correlation between QT interval and HR was only observed in dogs treated with timolol. In conclusion, levobunolol was the only drug that induced significant alterations in PS. Apraclonidine was the only drug that induced systemic hypotension. Timolol was the only drug to that induced a negative correlation between QT and HR.(AU)

O objetivo deste estudo foi avaliar as mudanças na pressão intraocular (PIO), no diâmetro pupilar (DP), na pressão sanguínea (PS), na frequência cardíaca (FC) e nas variáveis eletrocardiográficas (onda Pms, PmV, intervalo PR, complexo QRS, onda RmV e intervalo QT), ao longo do tempo da instilação do timolol 0,5%, do levobunolol 0,5% e da apraclonidina 0,5% em cães clinicamente normais. Dez Beagles adultos compuseram o estudo. Valores basais foram mensurados às oito,, 14 e 20 horas, durante três dias consecutivos. Foi instituído um período de espera de 10 dias entre a administração de cada fármaco. Durante 15 dias consecutivos, um olho de cada animal recebeu uma gota de cada um deles, a intervalos de 12 horas (às sete e às 19 horas). Os parâmetros foram avaliados nos momentos acima referidos, nos dias três, seis, nove, 12 e 15. Os dados foram comparados estatisticamente empregando-se o teste de Bonferroni após análise de variância para medidas repetidas (P<0,05). Teste de Pearson foi utilizado para correlação entre o intervalo QT com a FC e a PS. Não se encontrou diminuição da PIO. Observou-se redução significativa do DP na quase totalidade dos animais que receberam levobunol, relativamente ao olho controle. Diminuição significativa da FC foi vista ao terceiro dia após a administração do levobunolol, enquanto apraclonidina induziu aumento no 15º dia. A pressão arterial foi reduzida em todos os momentos com a apraclonidina. Observou-se correlação negativa entre o intervalo QT e a FC apenas nos indivíduos tratados com o timolol. Em conclusão, levobunolol foi o único fármaco que induziu alterações significativas no DP. A apraclonidina foi o único fármaco que induziu hipotensão sistêmica significativa. O timolol foi o único a ensejar correlação negativa entre o intervalo QT e a FC.(AU)

Ocular Metabolism of Levobunolol: Historic and Emerging Metabolic Pathways.

Although ocular transport and delivery have been well studied, metabolism in the eye is not well documented, even for clinically available medications such as levobunolol, a potent and nonselective ß-adrenergic receptor antagonist. Recently, we reported an in vitro methodology that could be used to evaluate ocular metabolism across preclinical species and humans. The current investigation provides detailed in vitro ocular and liver metabolism of levobunolol in rat, rabbit, and human S9 fractions, including the formation of equipotent active metabolite, dihydrolevobunolol, with the help of high-resolution mass spectrometry. 11 of the 16 metabolites of levobunolol identified herein, including a direct acetyl conjugate of levobunolol observed in all ocular and liver fractions, have not been reported in the literature. The study documents the identification of six human ocular metabolites that have never been reported. The current investigation presents evidence for ocular and hepatic metabolism of levobunolol via non-cytochrome P450 pathways, which have not been comprehensively investigated to date. Our results indicated that rat liver S9 and human ocular S9 fractions formed the most metabolites. Furthermore, liver was a poor in vitro surrogate for eye, and rat and rabbit were poor surrogates for human in terms of the rate and extent of levobunolol metabolism.

Bradyarrhythmias secondary to topical levobunolol hydrochloride solution.

An 88-year-old man was admitted with fatigue, dizziness, and heart palpitations. Both the electrocardiogram and Holter confirmed the existence of sinus bradycardia and sinus arrest. One hour prior to the onset of symptoms, he received levobunolol hydrochloride solution topically. The levobunolol hydrochloride solution was discontinued and the bradycardia resolved. He was diagnosed as having intermittent sinus bradycardia and sinus arrest, induced by topical ß-blocker therapy. Levobunolol hydrochloride solution is an effective therapy for ocular hypertension, probably by reducing aqueous fluid production. However, it can induce cardiac side effects such as bradyarrhythmia and should be used with caution in elderly patients or patients with cardiac disease.

Effects of anti-glaucoma drugs on resistive index of the medial long posterior ciliary artery using color Doppler imaging in Beagle dogs.

Color Doppler imaging (CDI) was carried out to evaluate the effects of anti-glaucoma drugs on ophthalmic circulation using CDI-derived resistive index (RI) values. CDI was performed on nine Beagle dogs, and RI values were calculated for the medial long posterior ciliary artery before and after the administration of anti-glaucoma drugs. A significant increase in RI values was found after topical administration of levobunolol (p < 0.05) or dipivefrin (p < 0.05). Pilocarpine showed no effects on RI values after topical administration. The results suggest that some anti-glaucoma drugs could affect ophthalmic blood flow.

Effects of anti-glaucoma drugs on resistive index of the medial long posterior ciliary artery using color Doppler imaging in Beagle dogs

Color Doppler imaging (CDI) was carried out to evaluate the effects of anti-glaucoma drugs on ophthalmic circulation using CDI-derived resistive index (RI) values. CDI was performed on nine Beagle dogs, and RI values were calculated for the medial long posterior ciliary artery before and after the administration of anti-glaucoma drugs. A significant increase in RI values was found after topical administration of levobunolol (p < 0.05) or dipivefrin (p < 0.05). Pilocarpine showed no effects on RI values after topical administration. The results suggest that some anti-glaucoma drugs could affect ophthalmic blood flow.

Effects of topical 0.5% levobunolol alone or in association with 2% dorzolamide compared with a fixed combination of 0.5% timolol and 2% dorzolamide on intraocular pressure and heart rate in dogs without glaucoma.

The goal of glaucoma management is to reduce intraocular pressure (IOP) and maintain it at a level compatible with the health of the optic nerve. New therapies are constantly being sought. Topical instillation of levobunolol 0.5%, alone or with dorzolamide 2%, has a hypotensive effect on the IOP in healthy dogs, and levobunolol combined with dorzolamide produces a stronger hypotensive effect than the combination of timolol and dorzolamide. All animals tolerate these topical medications well with no signs of discomfort, and no ocular side effects have been observed. Levobunolol, alone or in combination with dorzolamide, induces bradycardia, as does timolol with dorzolamide.

Poly (epsilon-caprolactone) microparticles containing Levobunolol HCl prepared by a multiple emulsion (W/O/W) solvent evaporation technique: effects of some formulation parameters on microparticle characteristics.

The aim of this study was to prepare poly (epsilon-caprolactone) (PCL) microparticles of Levobunolol HC1 (L-HC1) for use as an anti-glaucomatous drug to the eye. The double emulsion (W/O/W) solvent evaporation technique was used for encapsulating L-HC1 as a hydrophilic drug. The study examined the impact of different factors including the pH and volume of the external aqueous phase, the concentration of polyvinylalcohol (PVA) and Pluronic F68 (PF68) used as stabilizers and drug/polymer ratios on the characteristics of the microparticles. Scanning electron microscopy (SEM) and differential scanning calorimetry (DSC) were used to identify the physical state of the drug and polymer. The zeta potential of the particles was also identified. Entrapment efficiency was found to be highest with a 0.5% PVA concentration and 100 mL volume of external aqueous phase at pH 12. The high efficiency was due to a reduction in the degree of drug ionization. The microparticles were spherical and appropriately sized for ophthalmic application. Drug release from the microparticles appears to consist of two components, with an initial rapid release followed by a slower stage. Drug release was slower when the microparticle was incorporated into the thermally reversible gel (Pluronic F127) in comparison to drug release from the free drug incorporated into the gel and drug release from the free microparticle.

Effects of topical levobunolol or fixed combination of dorzolamide-timolol or association of dorzolamide-levobunolol on intraocular pressure, pupil size, and heart rate in healthy cats / Efeitos da aplicação tópica do levobunolol, da combinação fixa de dorzolamidatimolol ou da associação de dorzolamida com levobunolol sobre a pressão intra-ocular, o diâmetro pupilar e a freqüência cardíaca em gatos saudáveis

The effects of topical levobunolol with the fixed combination of 2 percent dorzolamide-0.5 percent timolol and the association of 2 percent dorzolamide with 0.5 percent levobunolol on intraocular pressure (IOP), pupil size (PS), heart rate (HR), and conjunctival hyperemia in eighteen halthy cats were investigated and compared. IOP, PS, HR, and conjuntival hyperemia were daily recorded at three times (9a.m., 2p.m., and 6p.m.). Three groups were formed (n=6), and one eye of each animal was randomly selected and treated with topical levobunolol (L), or commercial combination of dorzolamide-timolol (DT), or the association of dorzolamide with levobunolol (DL). The first day (0) consisted of recording of baseline values. On the next four consecutive days, drugs were instilled at 8a.m. and 8p.m. and measurements were taken at the same times fore cited. Comparing with the baseline values, all evaluated parameters significantly decreased (P<0.001). Conjuntival hyperemia was not seen. Levobunolol significantly declined IOP, PS, and HR in normal cats, and showed a stronger effect in lowering HR, when compared to dorzolamide-timolol effect. No synergistic effect in IOP declining was noted when levobunolol dorzolamide was added to levobunolol.

Estudaram-se e compararam-se os efeitos do levobunolol, da combinação fixa de dorzolamida 2 por cento-timolol 0,5 por cento e da associação de dorzolamida 2 por cento com levobunolol 0,5 por cento sobre a pressão intra-ocular (PIO), o diâmetro pupilar (DP), a freqüência cardíaca (FC) e a hiperemia conjuntival em 18 gatos saudáveis. PIO, DP, FC e hiperemia conjuntival foram aferidos diariamente, em três horários distintos (9h, 14h e 18h). Três grupos foram formados (n=6) e um olho de cada animal recebeu, aleatoriamente, uma gota de levobunolol (L), ou a combinação comercial à base de dorzolamida-timolol (DT), ou a associação de dorzolamida com levobunolol (DL). Parâmetros basais foram aferidos no primeiro dia (dia 0). Nos quatro dias consecutivos, os fármacos foram instilados às 8h e 20h e os parâmetros aferidos nos mesmos horários. Todos os parâmetros decresceram significativamente em relação aos valores basais (P<0,001) e não se observou hiperemia conjuntival. O levobunolol reduziu significativamente a PIO, o DP e a FC e o foi o fármaco que mais reduziu a FC. Não se observou efeito sinérgico na redução da PIO quando a dorzolamida foi adicionada ao levobulol.

An in vivo confocal microscopy and impression cytology analysis of preserved and unpreserved levobunolol-induced conjunctival changes.

PURPOSE: To provide an in vivo confocal microscopy (IVCM) and impression cytology analysis of preserved-and unpreserved levobunolol-induced changes of conjunctival epithelium. METHODS: 27 eyes of 27 patients were consecutively randomized to receive preserved or unpreserved levobunolol; all patients had a recent diagnosis of primary open angle glaucoma (POAG) or ocular hypertension and were not previously treated with topical medications. IVCM and impression cytology were performed before and after six months of therapy. Goblet cells density and a conjunctival epithelium regularity index were considered in the IVCM analysis, whereas impression cytology specimens were graded and scored in accordance with Nelson's method. RESULTS: After six months of therapy, IVCM and impression cytology parameters showed significant differences with respect to baseline in both groups (p<0.001); significant differences were also found between the two groups (p<0.001). The IVCM analysis showed a goblet cells density reduction (61% and 17% from baseline, respectively in group 1 and 2) (p<0.001) and an higher index of epithelial regularity (p<0.001) in both groups; the impression cytology analysis showed an higher score in both groups (p<0.001). CONCLUSIONS: All the IVCM and impression cytology parameters correlated well with the conjunctival modifications induced by the topical therapy, suggesting the less toxicity of unpreserved drugs.

Membrane electrodes for determination of some beta-blocker drugs.

Five poly(vinyl chloride) (PVC) matrix membrane electrodes responsive to the beta-blockers atenolol (AT), bisoprolol fumarate (BI), timolol maleate (TI), and levobunolol HCl (LV) were developed and characterized. A precipitation-based technique with ammonium reineckate anion as an electroactive material in PVC matrix with AT, BI, TI, and LV cations was used for fabrication of Electrodes 1-4, respectively. Electrode 5 fabrication was based on precipitation of LV cation with tungstophosphate anion as an electroactive material. Fast and stable Nernstian responses at 1 x 10(-2)-1 x 10(-7) M for different beta-blockers over the pH range of 2-8 were found for these electrodes, which were evaluated according to International Union of Pure and Applied Chemistry recommendations. The method was successively applied for the determination of beta-blockers in their pharmaceutical formulations. Validation of the method according to quality assurance standards showed the suitability of the proposed electrodes for use in the quality control assessment of these drugs. The recoveries for the determination of the beta-blocker drugs by the 5 proposed selective electrodes were 100.1 +/- 0.7, 99.9 +/- 0.8, 100.0 +/-1.1, 100.5 +/- 1.1, and 100.6 +/- 0.7% for Sensors 1-5, respectively. Statistical comparison between the results obtained by this method and the official method of the drugs was performed and no significant difference was found.

Vasodilatory mechanism of levobunolol on vascular smooth muscle cells.

Topical application of levobunolol hydrochloride, a beta-adrenergic antagonist used for treatment of glaucoma, is reported to increase ocular blood flow. We studied the mechanism of levobunolol-induced vasodilation in arterial smooth muscle. The effects of levobunolol or other agents on isolated, pre-contracted rabbit ciliary artery were investigated using an isometric tension recording method. The effects of the same agents on intracellular free calcium ([Ca(2+)](i)) in cultured human aortic smooth muscle cells were also studied by fluorophotometry. Levobunolol relaxed ciliary artery rings that were pre-contracted with either high-K solution, 1microM histamine, 10microM phenylephrine, or 100nM endothelin-1. The relaxation induced by levobunolol was concentration-dependent over the range of 10microM to 0.3mM. Inhibition of endothelial nitric oxide synthase or denudation of the endothelium did not affect this relaxation. Histamine-induced contractions were inhibited by the histamine H(1) antagonist pyrilamine. Radioligand binding experiments showed that levobunolol did not bind to the H(1) receptor. Further, histamine induced transient contraction in Ca(2+)-free solution, and levobunolol inhibited this contraction by 74.6+/-11.0%. In cultured smooth muscle cells in the presence of extracellular Ca(2+), levobunolol significantly inhibited the histamine-induced elevation of [Ca(2+)](i). However, it did not inhibit the increase of [Ca(2+)](i) in histamine-stimulated cells incubated in Ca(2+)-free solution. These results indicate that levobunolol may relax rabbit ciliary artery by two different mechanisms. First, the relaxation could be due to the blockade of Ca(2+) entry through non-voltage-dependent Ca(2+) channels. Second, levobunolol may change the Ca(2+) sensitivity of vascular smooth muscle cells.

Cellular cytotoxicity of antiglaucoma drugs in cultured corneal endothelial cells.

In this study, the various antiglaucoma drugs including betaxolol, timolol, levobunolol, carteolol, brimonidine, dipivefrin, dorzolamide, brinzolamide, latanoprost, unoprostone, and pilocarpine were used to investigate the effects of cellular cytotoxicity in cultured bovine corneal endothelial cells. After exposure to the drugs in three dilutions, 1/100, 1/1,000, and 1/10,000, for 100 minutes, cells were estimated based on the release assay of lactate dehydrogenase (LDH) enzyme. It was found that cellular LDH was significantly released in the medium only at 1/100th dilution of betaxolol, brimonidine, dorzolamide, dipivefrin, latanoprost and unoprostone to 130%, 123%, 145%, 157%, 128% and 237%, respectively, compared with controls upon exposure to drugs for 100 minutes. Moreover, benzalkonium chloride preservative at the concentrations ranging from 0.001 to 0.00001 mg/mL did not affect cellular LDH release in bovine corneal endothelial cells. These results indicate that high concentrations of antiglaucoma drugs may induce cytotoxicity in corneal endothelial cells.