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Doxorubicin (DOX) has been extensively utilized in cancer treatment. However, DOX administration has adverse effects, such as cardiac injury. This study intends to analyze the expression of TGF, cytochrome c, and apoptosis on the cardiac histology of rats induced with doxorubicin, since the prevalence of cardiotoxicity remains an unpreventable problem due to a lack of understanding of the mechanism underlying the cardiotoxicity result. Vernonia amygdalina ethanol extract (VAEE) was produced by soaking dried Vernonia amygdalina leaves in ethanol. Rats were randomly divided into seven groups: K- (only given doxorubicin 15 mg/kgbw), KN (water saline), P100, P200, P400, P4600, and P800 (DOX 15 mg/kgbw + 100, 200, 400, 600, and 800 mg/kgbw extract); at the end of the study, rats were scarified, and blood was taken directly from the heart; the heart was then removed. TGF, cytochrome c, and apoptosis were stained using immunohistochemistry, whereas SOD, MDA, and GR concentration were evaluated using an ELISA kit. In conclusion, ethanol extract might protect the cardiotoxicity produced by doxorubicin by significantly reducing the expression of TGF, cytochrome c, and apoptosis in P600 and P800 compared to untreated control K- (p < 0.001). These findings suggest that Vernonia amygdalina may protect cardiac rats by reducing the apoptosis, TGF, and cytochrome c expression while not producing the doxorubicinol as doxorubicin metabolite. In the future, Vernonia amygdalina could be used as herbal preventive therapy for patient administered doxorubicin to reduce the incidence of cardiotoxicity.
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Cardiotoxicidade , Vernonia , Ratos , Animais , Cardiotoxicidade/tratamento farmacológico , Cardiotoxicidade/etiologia , Cardiotoxicidade/prevenção & controle , Citocromos c/metabolismo , Etanol/efeitos adversos , Fator de Crescimento Transformador beta/metabolismo , Doxorrubicina/efeitos adversos , Apoptose , Extratos Vegetais/farmacologia , Estresse OxidativoRESUMO
BACKGROUND: The alkylating agent cyclophosphamide is used in chemotherapy regimens for various type of cancer. However, cyclophosphamide may lead to toxic side effects on the bladder, namely hemorrhagic cystitis, which can cause hematuria, and, potentially, bladder cancer. These effects are caused by acrolein, a byproduct of cyclophosphamide metabolism. In this study, a method to quantify 3-hydroxypropyl mercapturic acid (3-HPMA) in urine was developed. 3-HPMA is a stable metabolite of acrolein that serves as biomarker of acrolein. METHODS: Urine samples were collected 4 hours after cyclophosphamide administration and analyzed to determine the risk of hematuria. 3-HPMA was analyzed by reverse-phase LC-MS/MS using a triple quadrupole electrospray ionization mass spectrometer in the positive-ion mode. The mobile phase was a 90:10 (vol/vol) mixture of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. Multiple reaction monitoring mode was used, with m/z 222.10 â 90.97 for 3-HPMA and 164.10 â 122.02 for the internal standard N-acetyl cysteine (NAC). Samples were prepared by acidification and dilution. RESULTS: The analytical method produced a linear response within the concentration range of 40-10,000 ng/mL. The method was validated in accordance with 2018 FDA guidelines and applied to quantify 3-HPMA in the urine of 40 patients with breast cancer. The measured concentrations ranged from 820.3 to 5596.1 ng/mg creatinine. Seven patients identified with hematuria had low 3-HPMA concentrations of 4445.824 ± 411.17 ng/mg creatinine, and 33 patients without hematuria had low 3-HPMA concentrations of 2419.4 ± 1171.8 ng/mg creatinine. CONCLUSIONS: The method was applicable for the quantification of 3-HPMA in human urine. Large variations in 3-HPMA concentrations were found in 40 patients with breast cancer treated with cyclophosphamide, with a significant difference (P < 0.05) observed between patients with hematuria and those without hematuria.
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Acetilcisteína/análogos & derivados , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/urina , Ciclofosfamida/toxicidade , Ciclofosfamida/uso terapêutico , Acetilcisteína/urina , Acroleína/metabolismo , Adulto , Alquilantes/metabolismo , Alquilantes/uso terapêutico , Alquilantes/toxicidade , Biomarcadores/urina , Cromatografia Líquida/métodos , Ciclofosfamida/metabolismo , Feminino , Hematúria/induzido quimicamente , Humanos , Pessoa de Meia-Idade , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas em Tandem/métodosRESUMO
Telmisartan is highly variable drug indicated for treatment of hypertension. This study aimed to compare the bioavailability of two 80 mg telmisartan tablets in healthy Indonesian subjects. A randomized, open-label, single-dose, three-sequence, three-way, reference-formulation-replicated crossover study was conducted under fasting period with two-week washout period. In this study, 31 Indonesian subjects were enrolled and 28 subjects were completed the study. Serial blood samples were collected up to 72 h following drug administration. Plasma concentrations of telmisartan were determined using high-performance liquid chromatography method with fluorescence detector. The pharmacokinetic parameters of AUC0- t , AUC0-∞, and C max were assessed for bioequivalence. Bioequivalence acceptance was based on predefined criteria of 90% confidence interval (CI) of 80.00-125.00% for AUC parameters and reference-scaled-average bioequivalence of 71.73-139.42% for C max. The 90% CI for AUC0- t , AUC0-∞, and C max was 96.11-107.25%, 93.06-104.36%, and 94.23-127.01%, respectively. These results indicated that the two formulations of telmisartan were bioequivalent.
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Telmisartan , Administração Oral , Área Sob a Curva , Disponibilidade Biológica , Estudos Cross-Over , Humanos , Indonésia , Comprimidos , Equivalência TerapêuticaRESUMO
AIM: To compare the bioequivalence of two 10-mg memantine tablet formulations. MATERIALS AND METHODS: 19 subjects were included in this single-dose, open-label, randomized, two-way crossover study following an overnight fast. A 5-week washout period was applied. Blood samples were collected up to 72 hours following drug administrations. Plasma memantine concentrations were determined by LC-MS/MS. The pharmacokinetic parameters AUC0-72h and Cmax were calculated and used for bioequivalence evaluation after log-transformation. RESULTS: The point estimates and 90% confidence intervals for AUC0-72h and CCmax for memantine were 100.72% (97.43 - 104.13%) and 101.46% (97.15 - 105.96%), respectively. CONCLUSION: These results indicated that the two formulations of memantine were bioequivalent; therefore, they may be prescribed interchangeably.
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Memantina/farmacologia , Área Sob a Curva , Cromatografia Líquida , Estudos Cross-Over , Humanos , Indonésia , Comprimidos , Espectrometria de Massas em Tandem , Equivalência TerapêuticaRESUMO
Beraprost sodium is an oral prostacyclin analog that was first approved in 1992 (Japan) for the treatment of peripheral vascular disorders. It is administered orally as a tablet available in strength 20 µg. In this paper, we described a liquid chromatography tandem mass spectrometry method that was developed for the quantification of beraprost in human plasma with high sensitivity at picogram per milliliter concentration. The method had been validated in terms of selectivity, sensitivity, accuracy and precision, matrix effect, linearity, recovery and carry-over according to the Guideline on Bioanalytical Validation from the European Medicines Agency. The standard calibration curve for beraprost was 9.5-1419 pg/mL. This method has been applied successfully to a bioequivalence study with 60 µg of beraprost (three tablets) in 29 healthy volunteers. The results showed that the two formulations of beraprost are bioequivalent.
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Cromatografia Líquida/métodos , Epoprostenol/análogos & derivados , Espectrometria de Massas em Tandem/métodos , Epoprostenol/sangue , Epoprostenol/química , Epoprostenol/farmacocinética , Humanos , Modelos Lineares , Inibidores da Agregação Plaquetária/sangue , Inibidores da Agregação Plaquetária/química , Inibidores da Agregação Plaquetária/farmacocinética , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Equivalência TerapêuticaRESUMO
AIM: This study was conducted in order to compare the bioavailability of two film-coated tablets containing 25 mg of quetiapine. METHODS: 24 subjects were enrolled in and completed a single-center, randomized, single-dose, open-label, two-way crossover study with a 1-week washout period. Plasma samples were collected up to 24 hours following drug administration; thus, quetiapine was determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) method with turbo-ion-spray mode. The pharmacokinetic parameters used for bioequivalence assessment were AUC0-t, AUC0-∞, and Cmax. The 90% confidence intervals were obtained by analysis of variance for AUC0-t, AUC0-∞, and Cmax. RESULTS: The results were all within the range of 80.00 - 125.00%. CONCLUSION: Bioequivalence between formulations was concluded both in terms of rate and extent of absorption.â©.
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Antipsicóticos/farmacocinética , Fumarato de Quetiapina/farmacocinética , Adolescente , Adulto , Cromatografia Líquida , Estudos Cross-Over , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Comprimidos , Espectrometria de Massas em Tandem , Equivalência Terapêutica , Adulto JovemRESUMO
AIM: To compare the bioavailability of two 40-mg Rosuvastatin tablet formulations. METHODS: 24 subjects were included in this single-dose, open-label, randomized, two-way crossover study following an overnight fast. A 2-week wash out period was applied. Blood samples were drawn up to 72 hours following drug administrations. Rosuvastatin plasma concentrations were determined by liquid chromatography-tandem mass spectrometry method with TurboIon-Spray mode. Pharmacokinetic parameters AUC(0-t), AUC(0-∞), and Cmax were determined and used for bioequivalence evaluation after log-transformation, whereas tmax ratios were evaluated nonparametrically. RESULTS: The estimated point and 90% confidence intervals (CI) for AUC(0-t), AUC(0-∞), and C(max) for rosuvastatin were 95.21% (87.56 - 103.53%), 95.76% (88.01 - 104.18%), and 99.33% (89.37 - 110.41%), respectively. CONCLUSION: These results indicated that the two formulations of rosuvastatin were bioequivalent; therefore, they may be prescribed interchangeably.
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Inibidores de Hidroximetilglutaril-CoA Redutases/farmacocinética , Rosuvastatina Cálcica/farmacocinética , Área Sob a Curva , Química Farmacêutica , Estudos Cross-Over , Feminino , Humanos , Masculino , Comprimidos , Equivalência TerapêuticaRESUMO
Phenytoin is a first-line antiepileptic drug with narrow therapeutic range and follows non-linear pharmacokinetics. Pharmacokinetics of phenytoin have been studied in plasma matrix before, however, there were several disadvantages. This study aimed to obtain partial validation data of the analytical method and the pharmacokinetic profile of phenytoin in Dried Blood Spot (DBS) of six healthy subjects. DBS has the advantage of only requiring small sample volumes and could be transported more efficiently. Phenytoin along with carbamazepine as the chosen internal standard was analyzed with a reversed-phase high performance-liquid chromatography system and a photodiode array detector at 205 nm. The results of partial validation, which evaluated the linearity, within-run accuracy, and precision, were within the criteria acceptance range. The pharmacokinetic profile showed that average AUC0-t was 83.81 ± 37.32 µg.h/mL and AUC0-∞ was 83.65 ± 38.89 µg.h/mL with an average ratio of 93%. Previous study quantifying phenytoin in the plasma matrix found the average AUC0-t was 39.41 ± 8.57 µg.h/mL and AUC0-∞ was 42.94 ± 9.55 µg.h/mL. Despite the difference between parameters of phenytoin analyzed in DBS and plasma matrices, the pharmacokinetic profiles obtained from both matrices were similar indicated by comparable concentration-time curves, thus, proving that DBS matrix can be used interchangeably with the plasma matrix as a more comfortable and effective alternative to phenytoin quantification in blood.
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Background: Ivermectin is a broad-spectrum anthelmintic used to control onchocerciasis from nematode parasites. As an anthelmintic, ivermectin is designed to have high levels in the gastrointestinal tract, so that the systemic intake is relatively low. Due to the very small concentration of ivermectin, a selective and sensitive approach is needed for the analysis of ivermectin in blood. Several methods have been developed using plasma and Dried Blood Spots, but there are still shortcomings due to hematocrit effects. Therefore, this study was conducted to establish a validated ivermectin analysis method with doramectin as the internal standard in using Ultra High-Performance Liquid Chromatography-Tandem Mass Spectrometry. Methods: Mass spectrometry equipped with triple quadrupole and positive electrospray ionization mode was used to conduct the analysis. For the biological matrix, whole blood was used by Volumetric Absorptive Microsampling and extracted using a protein precipitation technique with a combination of acetonitrile and methanol (1:1). VAMS has some advantages such as not being affected by hematocrit, requires a small and fixed volume of sample, also a more efficient sampling process. Results: The optimum conditions were achieved with an Acquity® UPLC BEH C18 column (1,7 µm; 2.1 × 100 mm); extracted-flow rate was 0,2 mL/min; mobile phase was 5 mM ammonium formate pH 3.00 and acetonitrile (10:90) with isocratic elution. Multiple Reaction Monitoring (MRM) detection by m/z values was 892.41 > 569.5 for ivermectin and 916,41 > 331,35 for doramectin. Conclusion: The method has been appropriately validated in compliance with the 2018 guidelines laid out by the US Food and Drug Administration. Resulting the minimum detection (LLOQ) was 1 ng/mL with a linear concentration range spanning from 1 to 150 ng/mL.
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5-Fluorouracil is an antimetabolite drug indicated for cancer treatment. Therapeutic drug monitoring of 5-Fluorouracil is necessary because 5-Fluorouracil has narrow therapeutic window and its concentration in blood is affected by individual conditions, like gene polymorphisms. Dried Blood Spot (DBS) is one of the biosampling methods used for therapeutic drug monitoring. Asides from reducing patients' discomfort, the use of DBS can increase 5-Fluorouracil stability by stopping the enzymes activity in blood. Therefore, this research developed a method to monitor 5-Fluorouracil levels in DBS using ultra-high performance liquid chromatography-tandem mass spectrometry. Sample preparation was carried out by extracting DBS using 2-Propanol: ethyl acetate (16:84). Reconstituted samples were analyzed using ultra high performance liquid chromatography equipped with Acquity® UPLC BEH C18 column (2.1 × 100 mm; 1.7 µm). The ionization process was carried out in negative electrospray ionization mode. Multiple Reaction Monitoring (MRM) values were set at m/z 128.97 > 41.82 for 5-Fluorouracil and 168.97 > 57.88 for propylthiouracil as the internal standard. Optimum analytical conditions were obtained with acetonitrile-ammonium acetate 1 mM (95:5) as mobile phase, flow rate of 0.15 mL/min, and column temperature of 40 °C. The lowest level of quantification obtained from this method was 0.1 µg/mL with a calibration curve range of 0.1 µg/mL-60 µg/mL. This method was proven to be valid according to the requirements set by the US Food and Drug Administration and the European Medicines Agency.
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Objective: This study aims to develop and validate bioanalytical method for quantifying warfarin in VAMS samples using liquid chromatography tandem mass spectrometry (LC-MS/MS), directly implementing the method to patients receiving warfarin therapy. Methods: The UPLC-MS/MS method was developed and optimized, with quercetin as the internal standard. Sample preparation was carried out using protein precipitation with methanol-acetonitrile (1:3 v/v). Results: Chromatographic separation was achieved using Acquity® UPLC BEH C18 column with 0.1 % formic acid-acetonitrile-methanol (30:69:1 v/v) as mobile phase, in isocratic elution. Multiple Reaction Monitoring (MRM) detection was done using m/z values of 307.10 â 161.06 for warfarin and 301.03 â 150.98 for quercetin as internal standard, using Electrospray Ionization (ESI) negative ion source. The clinical application of the bioanalytical method was carried out on 25 patients receiving warfarin therapy at Universitas Indonesia Hospital and warfarin levels were well within the calibration range from 6.05 to 431.39 ng/mL. Conclusion: A novel method has been developed to analyze warfarin in VAMS samples. This method has been fully validated according to guideline from FDA 2022 and is linear in the range of 5-500 ng/mL and the value of r ≥ 0.9977, and successfully applied for the analysis of warfarin in VAMS samples of clinical patients.
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<b>Background and Objective:</b> Doxorubicin is an anticancer therapy belonging to the anthracycline class, which has clinical activity in breast cancer. Doxorubicin can cause cardiotoxic effects due to the formation of doxorubicinol as its main metabolite. The purpose of this study was to obtain the optimum sample preparation conditions for the analysis of doxorubicin in VAMS and as a form of therapeutic drug monitoring (TDM) in patients with cancer breasts. <b>Materials and Methods:</b> Analyze doxorubicin and doxorubicinol levels with Volumetric Absorptive Microsampling (VAMS) in patients' cancer breasts receiving doxorubicin in their therapeutic regimen. The sample was analyzed using Ultra Performance Liquid Chromatography tandem Mass Spectrometry (LC-MS/MS). The method uses deep linear range concentrations of 8-200 ng/mL for doxorubicin and 3-100 ng/mL for doxorubicinol. <b>Results:</b> Multiple reaction monitoring (MRM) value set at m/z 544.22>396.9 for doxorubicin; m/z 546.22>398.9 for doxorubicinol and m/z 528.5>362.95 for daunorubicin. The LLOQ value obtained was 8 ng/mL for doxorubicin and 3 ng/mL for doxorubicinol with linearity of 0.9904 for doxorubicin and 0.9902 for doxorubicinol. Analysis results show doxorubicin levels were in the range of 9.47 ng/mL to 87.84 ng/mL and doxorubicinol range between 4.24 and 54.02 ng/mL. <b>Conclusion:</b> Dosage cumulative doxorubicin ranges between 47.93 and 346.09 mg/m<sup>2</sup>; with this, the risk of cardiomyopathy in the patients surveyed is under 4%, according to the literature.
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Neoplasias da Mama , Cardiotoxicidade , Doxorrubicina , Monitoramento de Medicamentos , Feminino , Humanos , Antibióticos Antineoplásicos/efeitos adversos , Neoplasias da Mama/tratamento farmacológico , Cardiotoxicidade/etiologia , Doxorrubicina/efeitos adversos , Doxorrubicina/análogos & derivados , Monitoramento de Medicamentos/métodos , Espectrometria de Massa com Cromatografia Líquida , Espectrometria de Massas em TandemRESUMO
Favipiravir is a prodrug of T-1105 made by modifying the pyrazine group as a COVID-19 therapy. During the pandemic, a safe and comfortable biosampling technique is needed for the subject or patient. Volumetric Absorptive Microsampling (VAMS) is a biosampling technique with a small blood volume and minimum hematocrit effect. The aims of this study were to develop and validate an analytical method for quantifying favipiravir extracted from VAMS using High Performance Liquid Chromatography - Photodiode Array with remdesivir as an internal standard. Analysis of favipiravir was performed using a C18 column (Waters, Sunfire™ 5 µm; 250 × 4.6 mm), with injection volume of 50 µL, flow rate of 0.8 mL/min, column temperature 30 â, and wavelength 300 nm. The separation was conducted under gradient elution with mobile phase consists of acetonitrile-0.2 % formic acid-20 mM sodium dihydrogen phosphate pH 3.5 and run time 12 min. Sample preparation was carried out using a protein precipitation method with 500 µL of methanol as precipitating agent. Samples were mixed on vortex for 30 s, sonicated for 15 min, and centrifuged at 10,000 rpm for 10 min. Lower Limit of Quantification (LLOQ) obtained was 0.5 µg/mL and the calibration curve ranged from 0.5 to 160 µg/mL. Sensitivity, linearity, selectivity, carry-over, accuracy, precision, recovery, and stability were validated by the guideline from Food and Drug Administration 2018. The method developed has successfully met the full validation requirements by FDA 2018 with the LLOQ obtained was 0.5 µg /mL.
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COVID-19 , Humanos , Cromatografia Líquida de Alta Pressão/métodos , Cromatografia Líquida/métodos , PirazinasRESUMO
Favipiravir and remdesivir are drugs to treat COVID-19. This study aims to find an optimum and validated method for simultaneous analysis of favipiravir and remdesivir in Volumetric Absorptive Microsampling (VAMS) by Ultra High-Performance Liquid Chromatography-Tandem Mass Spectrophotometry. The use of VAMS can be an advantage because the volume of blood is small and the sample preparation process is simple. Sample preparation was done by precipitation of protein using 500 µL of methanol. Analysis was carried out by ultra high-performance liquid chromatography-tandem mass spectrophotometry with ESI+ and MRM with m/z 157.9 > 112.92 for favipiravir, 603.09 > 200.005 for remdesivir, and at m/z 225.968 > 151.991 for acyclovir as the internal standard. The separation was carried out using an Acquity UPLC BEH C18 column (100 × 2.1 mm; 1.7 m), 0.2% formic acid-acetonitrile (50:50), flow rate was 0.15 mL/min, and column temperature was 50°C. The analytical method has been validated with the requirements issued by the Food and Drug Administration (2018) and European Medicine Agency (2011). The calibration range of favipiravir is 0.5-160 µg/mL and 0.002-8 µg/mL for remdesivir.
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Lidocaine hydrochloride (LiH), an amide-type local anesthetic agent, is commonly used in dermatological procedures. LiH is categorized as a BCS (biopharmaceutics classification system) class III group, which has high solubility and poor permeability. It should be noted that, in this context, LiH is intended as a local anesthetic, so the level of LiH in systemic circulation should be minimized to avoid toxicity and unwanted side effects such as hypotension and bradycardia. This study aimed to formulate and evaluate LiH-loaded dissolving microneedles (DMNs) with different polymer bases. Moreover, an in vitro permeation study using Franz diffusion cells and in vivo study were also performed. LiH-loaded DMNs were prepared using polymer groups of poly(vinyl pyrrolidone) (PVP-K30) and hyaluronic acid (HA). DMNs were created using the micro-molding method with centrifugation. The formulations selected based on the evaluation were F3 (HA 10%) and F5 (PVP-K30 25%). Based on the in vitro permeation study, the amount of drug permeated and deposited in the skin at F3 (HA 10%) was 247.1 ± 41.85 and 98.35 ± 12.86 µg, respectively. On the other hand, the amount of drug permeated and deposited in the skin at F5 (PVP-K30 25%) was 277.7 ± 55.88 and 59.46 ± 9.25 µg, respectively. Our in vivo drug-permeation study showed that only one rat from the PVP-K30 polymer group-with a concentration of 150.32 ng/mL-was detected on rat plasma. Therefore, LiH can be formulated into a DMN and can be deposited in the skin with a safe concentration of the drug permeating into systemic circulation.
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Ketoprofen is an anti-inflammatory agent that may cause gastric irritation if administered orally. Dissolving microneedles (DMN) can be a promising strategy to overcome this issue. However, ketoprofen has a low solubility; therefore, it is essential to enhance its solubility using certain methods, namely nanosuspension (NS) and co-grinding (CG). This research aimed to formulate DMN containing ketoprofen-loaded NS and CG. Ketoprofen NS was formulated with poly(vinyl alcohol) (PVA) at concentrations of 0.5%, 1%, and 2%. CG was prepared by grinding ketoprofen with PVA or poly(vinyl pyrrolidone) (PVP) at different drug-polymer ratios. The manufactured ketoprofen-loaded NS and CG were evaluated in terms of their dissolution profile. The most promising formulation from each system was then formulated into microneedles (MNs). The fabricated MNs were assessed in terms of their physical and chemical properties. An in vitro permeation study using Franz diffusion cells was also carried out. The most promising MN-NS and MN-CG formulations were F4-MN-NS (PVA 5%-PVP 10%), F5-MN-NS (PVA 5%-PVP 15%), F8-MN-CG (PVA 5%-PVP 15%), and F11-MN-CG (PVA 7.5%-PVP 15%), respectively. The cumulative amounts of drug permeated after 24 h for F5-MN-NS and F11-MN-CG were 3.88 ± 0.46 µg and 8.73 ± 1.40 µg, respectively. In conclusion, the combination of DMN with nanosuspension or a co-grinding system may be a promising strategy for delivering ketoprofen transdermally.
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Cyclophosphamide (CP) is an anti-cancer alkylating prodrug, metabolized by CYP450 into its active metabolite 4-hydroxycyclophosphamide (4-OHCP). Its therapeutic effectiveness is determined by the 4-OHCP concentration. Several analytical methods in plasma and dried blood spots have been developed to analyze cyclophosphamide and 4-OHCP; however, there are many disadvantages. The objective of this study was to develop and validate the ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method by volumetric absorptive microsampling (VAMS) and 4-hydroxycyclophosphamide-d4 (4-OHCP-d4) as an internal standard. VAMS requires small sample volumes, and it is not affected by the hematocrit values; therefore, it is an efficient sampling method. The samples were derivatized with 5 µL semicarbazide hydrochloride (SCZ) and 25 µL of the resulting 4-OHCP-SCZ; 4-OHCP-d4-SCZ derivatives were absorbed by VAMS and extracted by protein precipitation. The optimum conditions were obtained using the Waters Acquity® UPLC BEH C18 (2.1 × 100 mm; 1.7 µm) column; flow rate 0.15 ml/min; mobile phase 0.01% formic acid and methanol; gradient elution mode for 6 min by positive electrospray ionization; and multiple reaction monitoring of m/z 260.7 > 140.0 for CP, 333.7 > 221.0 for 4-OHCP-SCZ, and 337.7 > 225.1 for 4-OHCP-d4-SCZ. The method met the validation requirements set by the FDA. The cyclophosphamide LLOQ value was 5 ng/mL, and the calibration curve range was 5-60,000 ng/ml. Furthermore, the 4-OHCP LLOQ value was 2.5 ng/ml, and the calibration curve range was 2.5-1,000 ng/ml.
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Gelatin is used as an additive in medicine, food, and cosmetics. Gelatin from goatskin is a new excipient that has not been explored by researchers, including for hard-shell capsules. The aim of this study was to evaluate and characterize the hard-shell capsules produced from goatskin gelatin. The goatskin gelatin was extracted by an acid hydrolysis method, and the functional properties were investigated. Hard-shell capsules were then produced from goatskin gelatin, evaluated, and characterized. The gelatin extracted from goatskin had 56.9% ± 0.95 clarity and a pH of 5.11 ± 0.09, 97.51% ± 1.1 protein content, 9.23% ± 0.08 water content, 0.18% ± 0.07 ash content, 2.08% ± 0.35 fat content, gel strength of 298 ± 2.64 gbloom, and viscosity of 27.33 ± 2.07 mPs. The gelatin has met the requirements to be made into hard-shell capsules. The average weight of the hard-shell capsules produced was 96.9 mg with 8.69 standard deviation. The average size of the body and cap length was 18.84 ± 0.64 mm and 10.98 ± 0.30 mm, respectively. The results of capsule evaluation and characterization were as follows: the pH was 4.82 ± 1,27, water content was 10.03 ± 0.21, disintegration time was 4.02 ± 2.09 min, and there was no microbial growth. Thus, the capsules made have met the requirements and can be produced in a large quantity.
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N7-(2-carbamoyl-2-hydroxyethyl)guanine (N7-CAG) is a DNA adduct formed by glycidamide, which is the metabolite of acrylamide. Acrylamide can be found in foods containing reducing sugars and asparagine that are heated at high temperatures. Analysis of N7-CAG was performed in Dried Blood Spot (DBS) samples from 25 subjects of group test who consumed a lot of acrylamide-containing foods and 25 subjects of negative control group. This study aimed to determine whether there is a significant difference in the levels of N7-CAG between the two groups. DBS samples were extracted using the QIAamp DNA Mini Blood Kit and analyzed using Ultra High Performance Liquid Chromatography-Tandem Mass Spectrometry (UHPLC-MS/MS). Separation was performed using an Acquity UPLC BEH C18 column (2.1 mm × 100 mm; 1.7 µm), eluted a flow rate of 0.1 ml/min under an isocratic of mobile phase of 0.1% formic acid and acetonitrile. The bioanalytical method of N7-CAG in DBS with allopurinol as the internal standard by using UHPLC-MS/MS has been validated. The calibration curve range of N7-CAG obtained was 10-300 ng/ml with a coefficient of correlation of 0.997. The results of the analysis on 25 test group subjects showed that the concentration of N7-CAG ranged from 1.87 to 23.71 ng/ml, while the 25 subjects in the negative group ranged from 1.18 to 8.47 ng/ml. The results of the Mann Whitney test showed that there was a significant difference in the levels of N7-CAG between the test group and the negative control group with p value less than 0.001.
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Intranasal drug transport through the olfactory route to the brain is an effective drug route for increased absorption and bioavailability of the drug. The objective of this study was to increase the penetration of valproic acid as an anticonvulsant into a delivery system comprising liposomes. Valproic acid liposomes were prepared by a thin-layer hydration technique using soybean phosphatidylcholine and cholesterol as the main ingredients. The formulations were evaluated for diameter size, entrapment efficiency (EE), zeta potential, polydispersity index, and morphology. ex vivo permeation using sheep nasal mucosa and in vivo efficacy were assessed by performing a pharmacokinetic study in Wistar albino rats following intranasal administration of the formulations in comparison with pure drug. The mean size particle of optimized liposomes ranged from 90 to 210 nm with a low polydispersity index (<0.5). The EE of optimized liposomes was between 60% and 85%, increasing the concentration of phosphatidylcholine added to the formula. Transmission electron microscopy observations (40,000×) showed that valproic acid liposomes have a spherical molecular shape and a particle size of below 250 nm. The ex vivo and in vivo results showed that liposomal formulations provided enhanced brain exposure. Among the formulations studied, Formula 4 (F4) showed greater uptake of valproic acid into the brain than plasma. The high brain targeting efficiency index for F4 indicated the preferential transport of the drug to the brain. The study demonstrated the successful formulation of surface-modified valproic acid liposomes for nasal delivery with brain targeting potential.