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Abstract Introduction. A specialized service for antifungal blood level determination is not available in Colombia. This service is essential for the proper follow-up of antifungal therapies. Objective. To standardize and validate a simple, sensitive, and specific protocol based on high-performance liquid chromatography with a diode array detector for voriconazole blood level quantification. Materials and methods. We used an Agilent HPLC™ series-1200 equipment with a UV- diode array detector with an analytical column Eclipse XDB-C18 and pre-column Eclipse- XDB-C18 (Agilent). We used voriconazole as the primary control and posaconazole as an internal control. We performed the validation following the Food and Drug Administration (FDA) recommendations. Results. The best chromatographic conditions were: Column temperature of 25°C, UV variable wavelength detection at 256 nm for voriconazole and 261 nm for posaconazole (internal standard); 50 μl of injection volume, 0,8 ml/min volume flow, 10 minutes of run time, and mobile phase of acetonitrile:water (60:40). Finally, retention times were 3.13 for voriconazole and 5.16 minutes for posaconazole. Quantification range varied from 0.125 μg/ml to 16 μg/ml. Conclusion. The selectivity and chromatographic purity of the obtained signal, the detection limits, and the standardized quantification make this method an excellent tool for the therapeutic monitoring of patients treated with voriconazole.
Resumen Introducción. Hasta la fecha, Colombia no cuenta con un servicio especializado de medición de niveles séricos de antifúngicos, procedimiento esencial para el adecuado seguimiento del tratamiento de infecciones fúngicas invasoras. Objetivo. Estandarizar y validar un protocolo -simple, sensible y específico- basado en la aplicación de cromatografía líquida de alta eficiencia acoplada con un detector de arreglo de diodos para la cuantificación de los niveles séricos de voriconazol. Materiales y métodos. Se usó un equipo HPLC-AgilentTM, serie-1200, con un detector UVDAD, una columna analítica Eclipse-XDB-C18 y una pre-columna Eclipse-XDB-C18, ambas de la marca Agilent. Como control primario se utilizó voriconazol y como control interno, posaconazol. La validación se hizo cumpliendo todos los criterios de aceptación recomendados por la Food and Drug Administration (FDA). Resultados. Las mejores condiciones cromatográficas se obtuvieron con los siguientes parámetros: temperatura de la columna de 25 °C, detección UV-VWD de 261 nm, volumen de inyección de 50 μl, flujo de 0,8 ml/minuto y un tiempo de corrido de 10 minutos. La fase móvil usada fue acetonitrilo:agua (60:40) y los tiempos finales de retención fueron de 3,13 para voriconazol y de 5,16 minutos para posaconazol. El rango de cuantificación fue desde 0,125 μg/ml hasta 16 μg/ml. Conclusiones. La selectividad y la pureza de la señal cromatográfica, así como los límites de detección y cuantificación estandarizados hacen de esta metodología una excelente herramienta para el seguimiento terapéutico de pacientes tratados con voriconazol o en profilaxis con este fármaco.
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Objective:A high performance liquid chromatography (HPLC) method was developed to determine the enzymatic activity of CD38 in blood, which was the major enzyme responsible for consuming nicotinamide adenine dinucleotide (NAD). Additionally, the study aimed to detect the differences in CD38 enzymatic activity among individuals of varying ages and health statuses.Methods:A 50 μl whole blood matrix and enzyme reaction substrate of 150 μl β-NAD at a concentration of 500 μmol/L were selected for the analysis. To eliminate the impact of endogenous β-NAD, the whole blood sample was pre-incubated at 37 ℃ for 20 minutes before adding the substrate. The reaction was terminated by perchloric acid (PCA) after incubation at 37 ℃ for 40 min. The change in product nicotinamide (NAM) before and after the enzymatic reaction was measured by HPLC to calculate the CD38 activity. The linearity, limit of detection, limit of quantification, precision, and stability of the method were evaluated. The CD38 enzymatic activities in 60 healthy volunteers and 30 colorectal cancer patients in blood were determined by the developed method.Results:Pre-incubation at 37 ℃ for 20 minutes eliminated the effect of endogenous β-NAD. The correlation coefficient of NAM was 0.999 in the concentration range of 0.1-3.2 μmol/L, with limit of detection of 0.5 nmol/L and limit of quantification of 2.1 nmol/L. The average within-run imprecision ( CV) and total CV were 3.22%-4.03% and 2.91%-4.70%, respectively. The recovery rate ranged from 94.82% to 96.81%. The CD38 activity of whole blood was stable by storage at 4 ℃ for 48 hours, storage at room temperature for 8 hours, thawing of frozen whole blood at room temperature for 2 hours, or repeated freeze-thawing three times. NAM, NAD standards, and pre-treatment samples were stable after 48 hours at 4 ℃ and 8 hours at room temperature. CD38 activity gradually decreased with increasing concentration of the added CD38 inhibitor 4-aminoquinoline derivative (78c). Measurement of 60 healthy physical examination population samples showed significantly higher CD38 enzyme activity in the elderly group than that in the young group ( t=-2.776, P=0.007) and measurement of 30 colorectal cancer patients showed significantly higher CD38 enzyme activity than that in healthy people ( t=-2.572, P=0.012). Conclusion:The established HPLC method for determining CD38 enzymatic activity is characterized by its simplicity, efficiency, accuracy, and reproducibility. This technique serves as a valuable tool for investigating aging and aging-related diseases.
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Objective:To establish reverse-phase high-performance liquid chromatography and simultaneously determine gallic acid, methyl gallate, corilagin, Sennoside B, and Sennoside A levels in Sana preparations.Methods:From January to December 2021, Phenomenex Hydro-RP 80A column (4.60 mm × 250 mm, 4 μm) was used. Elution was conducted using mobile phases methanol (A)-0.2% formic acid (B). The following gradients were applied: 1%-3%A for 0-18 minutes, 3%-15%A for 18-19 minutes, 15%-17%A for 19-40 minutes, 17%-25%A for 40-45 minutes, 25%-35%A for 45-65 minutes, 35%-60%A for 65-95 minutes, 60%-90%A for 95-96 minutes, 90%-1%A for 96-97 minutes. The flow rate was 1.0 mL/minute. The column temperature was 35 ℃. The detection wavelength was 270 nm.Results:The linear ranges of gallic acid, methyl gallate, corilagin, Sennoside B and Sennoside A were 0.182-1.099 μg ( r = 0.999 9), 0.046-0.278 μg ( r = 0.999 2), 0.266-1.598 μg ( r = 0.999 4), 0.172-1.036 μg ( r = 0.999 2), and 0.176-1.056 μg ( r = 0.999 9). The average dosing recovery rates were 100.02%, 99.14%, 99.38%, 101.77%, and 100.92%, respectively. Conclusion:Reverse-phase high-performance liquid chromatography can be used for quality control of Sana preparations because of high accuracy, sensitivity, reliability, and reproduction.
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Abstract Introduction: Various methods of analysis for the assay of chemotherapeutic agent 5-Fluorouracil (5-FU) in human and animal biological fluids have previously been reported. However, there is no standardization for detecting 5-FU in the hamsters' saliva that received the chemotherapeutic agent. Objective: Considering that the administration of 5-FU in some way changes the morphology and function of the salivary glands, and that the presence of the chemotherapeutic agents in the oral mucosa may lead to some oral complications, the aim of this study was to determine the presence of 5-FU in the hamsters' saliva that received the chemotherapeutic agent, by means of the High Performance Liquid Chromatography technique (HPCL) since this animal model is used in studies of 5-FU induced oral mucositis and glandular hypofunction. Methods: Twelve animals were divided into 4 groups: CP and CPI, in which the animals received pilocarpine (CP) or pilocarpine + isoproterenol (CPI) and the chemotherapy vehicle intraperitoneally; and Groups QP and QPI, in which the animals received the same secretagogues listed above, and the chemotherapeutic agent 5-FU, respectively. After the secretagogue administration, saliva was collected from all the animals for a period of 60 mins. Subsequently, the saliva was frozen at -80 ËC for later determination of the chemotherapeutic agent by HPLC. After the the chromatograms analysis, and based on the results obtained, it was possible to identify the presence of 5-FU in the saliva samples from hamsters that received the chemotherapeutic agent intraperitonally, by the HPLC technique. (AU)
Resumo Vários métodos de análise para o ensaio do quimioterápico 5-Fluorouracil (5-FU) em fluidos biológicos de humanos e animais, foram previamente relatados. No entanto, não há uma padronização para detecção de 5-FU na saliva de hamsters que receberam o quimioterápico. Considerando que a administração do 5-FU altera de alguma maneira a morfologia e função das glândulas salivares, e que a presença do quimioterápico na mucosa oral pode levar a algumas complicações orais, este trabalho teve como objetivo de determinar a presença de 5-FU na saliva de hamsters que receberam o quimioterápico pela técnica de Cromatografia Líquida de Alta Eficiência (CLAE), uma vez que este modelo animal é usado nos estudos com mucosite oral e hipofunção glandular, induzidas por 5-FU. Doze animais foram divididos em 4 grupos: CP e CPI, onde os animais receberam intraperitonealmente pilocarpina (CP) ou pilocarpina + isoproterenol (CPI) e o veículo do quimioterápico, e os grupos QP e QPI, onde os animais receberam, respectivamente, os mesmos secretagogos listados acima e o quimioterápico 5-FU. Após a administração do secretagogo, foi coletada a saliva de todos os animais, por um período de 60 min. Em seguida, a saliva foi congelada a -80 ËC para posterior determinação do quimioterápico por CLAE. Após análise dos cromatogramas, e com base nos resultados obtidos, foi possível identificar a presença do 5-FU nas amostras de saliva de hamsters que receberam o quimioterápico via intraperitoneal pela técnica da CLAE. (AU)
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Objective:To explore the relation of the radiochemical purity and in vivo imaging effect of 68Ga-1, 4, 7, 10-tetraazacyclododecane-1, 4, 7, 10-tetraacetic acid (DOTA)- D-phe1-Tyr3-Thr8-octreotide (TATE) injection. Methods:High performance liquid chromatography (HPLC) and thin-layer chromatography (TLC) methods were established to determine 68Ga-DOTATATE, 68Ga 3+ , 68Ga in colloidal form and 68Ga-DOTA- D-Phe1-Tyr3-Thr8-dethreonine-octreotide (heptapeptide) and to study the influence of precursor purity on radiochemical purity of labelled products. The uptake of 68Ga-DOTATATE injection with different radiochemical purities was investigated in nude mice bearing AR42J cells by microPET imaging and the tumor target/non-target (T/NT) value was calculated. One-way analysis of variance and Pearson correlation analysis were used to analyze the data. Results:The contents of 68Ga 3+ and 68Ga in colloidal form were not related with precursor purity ( r values: 0.385, 0.497, P values: 0.306, 0.137), while the content of 68Ga-DOTA-heptapeptide was positively related with the purity of DOTA-heptapeptide ( r=0.957, P<0.001). The radiochemical purities of 68Ga-DOTATATE injection were (87.0±2.3)%, (86.8±0.8)% and (94.0±3.1)% when the DOTATATE purities were 90.9%, 91.6% and 99.2%, respectively. The results of microPET imaging showed that the tumor uptake was positively related with the radiochemical purity of 68Ga-DOTATATE injection ( r=0.828, P<0.001), and the T/NT values of 68Ga-DOTATATE injection with radiochemical purities of 95.7%, 85.8%, 84.5% and 79.9% were 21.25±8.84, 8.50±1.51, 11.38±1.65 and 6.01±0.99, respectively ( F=11.48, P=0.001). Conclusion:The radiochemical purity of 68Ga-DOTATATE injection is impacted by the purity of labelled precursor and manufacturing processes and is related with the imaging effect in vivo.
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Objective:To analyze the changes in serum metabolites of patients with uremia using ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS), and provide a theoretical basis for the prevention and treatment of uremia.Methods:Uremia patients from the Department of Nephrology, the Affiliated Hospital of Southwest Medical University, and the volunteers from the Health Examination Center were enrolled in this study. According to the inclusion and exclusion criteria, 20 uremia patients (experimental group) and 20 volunteers (control group) were screened out. UHPLC-MS was used to detect the metabolites in the serum of subjects from the two groups, and difference analysis was made to screen the different metabolites, followed by correlation analysis and pathway enrichment study.Results:A total of 412 metabolites were identified by UHPLC-MS. Principal components analysis (PCA) proved that these metabolites could distinguish the control group and the experimental group well. The criteria [variable importance for the projection (VIP)>1, fold changes (FC)>1.25 or FC<0.8 and P value<0.05] was set to screen those significantly different metabolites. Finally, 28 significantly different metabolites were screened out, of which 18 metabolites increased significantly, the other 10 different metabolites decreased significantly. Correlation analysis results proved a certain correlation among 28 different metabolites and the experimental group and control group samples, and between the 28 differential metabolites themselves. Enrichment analysis found that 28 different metabolites might enrich the catecholamine biosynthetic pathway, and pathway analysis suggested that 28 different metabolites might affect glutamate, aspartame acid and glutamate metabolic pathways. Conclusion:Based on metabonomic analysis, some metabolites in the serum of patients with uremia have changed, which can affect some metabolic pathways, thus affecting the pathophysiological process of patients with uremia.
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Objective:To establish the TLC identification method and content determination method of ferulic acid, ligustilide, hydroxysafflor yellow A and paeoniflorin in Shangke Huoxue Decoction for quality evaluation.Methods:Ferulic acid, ligustilide, hydroxysafflor yellow A and paeoniflorin were qualitatively identified by TLC method, and the content was determined by HPLC method. Waters Symmetry ShieldTM RP18 column (4.6 mm×250 mm, 5 μm) was set, the mobile phase consisted of acetonitrile-0.15% phosphoric acid water with gradient elution at a flow of 1.0 ml/min, and the column temperature was 30 ℃.The detection wavelength was 320 nm (33-50 min for ferulic acid, 55-70 min for ligustilide), 403 nm (7-31 min for hydroxysafflor yellow A) and 230 nm (7-31 min for paeoniflorin).Results:The TLC spots were clear. The linear relationships of ferulic acid, ligustilide, and hydroxysafflor yellow A were good in the range of 3.05-48.74 μg, 3.50-26.24 μg, 21.34-213.44 μg. The method was stable, repeatable with good recovery rate.Conclusion:The TLC and HPLC method for the simutanous determination of the four effective components in Shangke Huoxue Decoction were established, and the methods are suitable for the quality evaluation of Shangke Huoxue Decoction.
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Objective:To establish a method for quantitative analysis of the active ingredients including salidroside, rosarin and rosavin and content determination in Rhodiola rosea at different harvest months. Methods:HPLC was used on an X selectHSS T3 (250 mm×4.6 mm, 5 μm) column with mobile phase consisting of methol-acetonitrile-phosphoric acid (0.05%) aqueous solution for gradient elution at a flow rate of 1 ml/min. The wavelength was detected at 275 nm (salidroside) and 254 nm (rosarin, rosavin). The column temperature was set at 30 ℃ and the injection volume was 5 μl.Results:The peak areas of Salidroside, rosarin and rosavin showed good linear relationships ( r > 0.999) with the content in the ranges of 44-1 420, 10-307 and 18-573 μg, respectively. The method was precise, stable, repeatable and the sample recovery test all well satisfied the requirements of quantitative analysis. The highest accumulation of the active ingredients was observed in Rhodiola rosea in September and the content of salidroside, rosarin and rosavin were 0.66, 0.07 and 0.53 mg/g, respectively. Conclusion:This method is simple and rapid to evaluate the content of active ingredients in Rhodiola rosea.
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ABSTRACT Hemoglobin A1c (HbA1c) measurement is commonly performed in diabetes mellitus patients to monitor glycemic control over the last three to four months. Carbamylated hemoglobin, which is the hemoglobin that binds to isocyanic acid derived from urea, is one of the possible analytical interference in the uremic patient. When measured by ion-exchange high-performance liquid chromatography (HPLC), carbamylated hemoglobin forms a peak that overlaps the peak of HbA1c, causing a falsely elevated HbA1c result. We report a case of a 60-years-old man who had a spurious increase in HbA1c, with a high carbamylated hemoglobin peak disproportionate to the urea value. Subsequent hemoglobin analysis using hemoglobin electrophoresis and HPLC hemoglobin testing system suggested hemoglobin J (Hb J) variant. Our case highlighted the possibility of misleading HbA1c interpretation in the presence of a high carbamylated hemoglobin peak, but not proportionate to urea value. In this study, Hb J was detected. A method free from hemoglobin variant interference should be used ideally, and monitoring glycemic control should be performed using alternative methods, such as serum fructosamine or continuous glucose monitoring.
RESUMEN La prueba de la hemoglobina glicosilada A1c (HbA1c) se realiza comúnmente en pacientes con diabetes mellitus para monitorear el control glucémico durante los últimos tres o cuatro meses. La hemoglobina carbamilada, que es la hemoglobina que se une al ácido isociánico derivado de la urea, es una de las posibles interferencias analíticas en el paciente urémico. Cuando se mide mediante cromatografía líquida de alta eficacia (HPLC) de intercambio iónico, la hemoglobina carbamilada forma un pico que se superpone al pico de HbA1c, lo que provoca un resultado de HbA1c falsamente elevado. Presentamos el caso de un hombre de 60 años que tuvo un aumento espurio de HbA1c, con un pico de hemoglobina carbamilada alto desproporcionado al valor de urea. El análisis de hemoglobina posterior mediante electroforesis de hemoglobina y el sistema de prueba de hemoglobina HPLC sugirió una variante de hemoglobina J (Hb J). Nuestro caso destacó la posibilidad de una interpretación engañosa de la HbA1c en presencia de un pico de hemoglobina carbamilada alto, pero no proporcional al valor de urea. En este estudio, se detectó Hb J. Lo ideal sería utilizar un método libre de interferencia de variantes de hemoglobina, y la monitorización del control glucémico debería realizarse mediante métodos alternativos, como la fructosamina sérica o la monitorización continua de la glucosa.
RESUMO A dosagem de hemoglobina A1c (HbA1c) é comumente realizada em pacientes com diabetes mellitus para monitorar o controle glicêmico nos últimos três a quatro meses. A hemoglobina carbamilada - hemoglobina que se liga ao ácido isociânico derivado da ureia - é uma das possíveis interferências analíticas no paciente urêmico. Quando medida por cromatografia líquida de alta eficiência (HPLC) por troca iônica, a hemoglobina carbamilada forma um pico que se sobrepõe ao pico da HbA1c, causando um resultado falsamente elevado da HbA1c. Relatamos o caso de um homem de 60 anos de idade que apresentava um aumento espúrio de HbA1c, com alto pico de hemoglobina carbamilada desproporcional ao valor de ureia. A análise subsequente da hemoglobina usando eletroforese de hemoglobina e sistema de teste de hemoglobina por HPLC sugeriu a variante de hemoglobina J (Hb J). Nosso caso destacou a possibilidade de interpretação enganosa da HbA1c na presença de um pico alto de hemoglobina carbamilada, mas não proporcional ao valor da ureia. Neste estudo, foi detectada a Hb J. Um método isento de interferência de variantes da hemoglobina deve ser idealmente usado, e o monitoramento do controle glicêmico deve ser feito usando métodos alternativos, como frutosamina sérica ou monitoramento contínuo da glicose.
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Resumen Justificación: La genética en las variantes de hemoglobina en Costa Rica es resultado del cruce de caracteres autóctonos indígenas con poblaciones inmigrantes de europeos, africanos y otros, desde el periodo de la Conquista, que contribuyeron a la mezcla genética presente en la población de Costa Rica. Las hemoglobinopatías mayormente distribuidas en la población humana son: hemoglobina S, C, D y E, siendo la hemoglobina S la más frecuente y la que presenta consecuencias más graves. Objetivo: Detectar variantes de hemoglobina en pacientes examinados por hemoglobina A1c, en la sección de Química Clínica del laboratorio de la Clínica de Filadelfia de la Caja Costarricense de Seguro Social, en el Cantón de Carrillo, Guanacaste, Costa Rica, durante el período de enero a octubre de 2018. Métodos: Se analizaron 2775 muestras sanguíneas de pacientes de los nueve equipos básicos de salud que conforman el Área de Salud de Carrillo, y que además requieren estudio por hemoglobina glicosilada en el período de enero a octubre de 2018. El análisis se realizó en el Laboratorio del Área de Salud de Carrillo. Las muestras fueron recolectadas en tubos vacutainer con EDTA y analizadas en el equipo automatizado TOSOH HLC-723GX, utilizando la metodología HPLC cromatografía de intercambio catiónico con la separación y cuantificación de las diferentes fracciones de hemoglobina. Los datos se analizaron en plantilla de Microsoft Excel. Resultados: En 2775 pacientes examinados por hemoglobina A1c, 167 (6,0 %) fueron portadores de variantes de hemoglobina, con una frecuencia de 1/17, en donde el 97 % correspondió a heterocigotos para hemoglobina un 3% a heterocigotos para hemoglobina C y ninguno para la variante D. La presencia de variantes se observó en los 9 equipos básicos de atención integral en salud del área. La distribución de portadores por equipo básico de atención en el Área de Salud de Carrillo varió de un 4,0 % a un 9,3 %. Conclusiones: Un 6 % de las muestras analizadas presentó variantes de hemoglobina, siendo la hemoglobina S la predominante. Esta característica presente en la población del cantón de Carrillo merece atención a nivel de salud pública; la metodología existente a nivel de área permite estudiar a un grupo de población (costo efectivo) en riesgo que precisa vigilancia y asesoramiento genético, con el fin de concienciar a la población respecto al problema, reducir la incidencia de la enfermedad y prolongar la supervivencia de los afectados.
Abstract Background: The genetics in hemoglobin variants in Costa Rica are a result of the crossing of autochthonous indigenous characters with European, African and other immigrant populations. All of these contributed to the genetic mixture that is currently present in Costa Rica's population. The most distributed hemoglobinopathies in the human population are: hemoglobin S, C, D, and E, with hemoglobin S being the most frequent and having the most serious consequences. Objective: Detection of hemoglobin variants in patients who were examined for hemoglobin A1c in the Clinical Chemistry section of the Filadelfia Clinic Laboratory, from January to October 2018. The clinic is in Canton of Carrillo, Guanacaste (Costa Rica), and it is part of the social security system. Methods: 2775 blood samples and their respective data were analyzed from patients of the nine basic health teams that make up the Carrillo Health Area and required a study for glycosylated hemoglobin from January to October 2018. The analysis was performed in the Carrillo Health Area Laboratory. The samples were collected in vacutainer tubes containing EDTA and analyzed in the TOSOH HLC-723GX automated equipment, using the HPLC cation exchange chromatography methodology with the separation and quantification of the different hemoglobin fractions. The data was then analyzed in a Microsoft Excel template. Results: In the 2775 patients examined for hemoglobin A1c, 167 (6.0%) were found to be carriers of hemoglobin variants, with a frequency of 1/17, where 97% corresponded to heterozygotes for hemoglobin S, 3% heterozygous for hemoglobin C, and none for variant D. The presence of variants was observed in the 9 basic teams of integral health care of the area, and the distribution varied from 4% to 9,3% between them. Conclusions: A total of 6% of the samples analyzed showed a hemoglobin variant, being hemoglobin S the most predominant. This characteristic present in the population of Canton of Carrillo deserves attention at the public health level. The existing methodology at the level area allows professionals to study a population group at risk that deserves surveillance and genetic counseling, in order to raise awareness about the problem, reduce the incidence of the disease, and prolong the survival of those affected by it.
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Humains , Hémoglobines/génétique , Costa Rica , Hémoglobinopathies , DrépanocytoseRÉSUMÉ
Objective To determine the contents of genistin and genistein in flemingia philippinensis by HPLC. Methods The contents of genistin and genistein in flemingia philippinensis were quantitatively determined by HPLC method. Results The linear relationship between genistin and genistein was good (r=0.9999, RSD=1.77％, 1.16％). There were differences in the contents of genistin and genistein in flemingia philippinensis of different regions, and the genistin content in Yangshuo city of Guangxi was the highest (0.575 mg/g), while the genistein content in Fuzhou city of Guangxi was the highest (0.264 mg/g). Conclusions The HPLC method has good reproducibility and stability in measuring the contents of genistin and genistein in flemingia philippinensis. The contents of genistin and genistein are different in flemingia philippinensis from different areas in Guangxi.
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Objective To establish an ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for detecting α-hydroxybutyrate (α-HB) in serum.Methods Electrospray ionization negative ion and multiple reaction monitoring mode were used to detect serum α-HB.The linearity,low limits of quantification,precision,recovery and interference of UHPLC-MS/MS were evaluated.The reference interval of this method was established in 130 serum samples (62 males and 68 females) from Shanghai East Hospital.Dixon method was used to judge the outliers and K-S test was used to analyze the data normality.The standard curve was scored by linear regression analysis.Results The total run time was 4 min of UHPLC-MS/MS method for the determination of α-HB.It has a good linear relationship in the range of 0.5-40.0 mg/L(r=0.999 4);the low limit of quantification was 0.5 mg/L;the in-batch and inter-batch coefficient of variation precision were less than 4.1％ and 6.3％,respectively;the recovery ranged between 95.8％ and 103.8％.Hemolytic samples (about 5 g/L hemoglobin),lipemic samples (about 12 mmol/L triglyceride),icteric samples (about 150 μmol/L total bilirubin) had no significant interference to the detection.The reference range of the apparent healthy population was 1.46-6.48 mg/L.Conclusions A method for the determination of serum α-HB by UHPLC-MS/MS was established.The method was simple,rapid,and could be used for the detection of clinical samples.
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A simple, sensitive, precise, accurate and robust high performance liquid chromatographic method has been developed for simultaneous estimation of saxagliptin (SAXA) and dapagliflozin (DAPA) in pharmaceutical formulation. Design of experiments (DoE) was applied for multivariate optimization of the experimental conditions of RP-HPLC method. Risk assessment was performed to identify the critical method parameters. Three independent factors; mobile phase composition, flow rate and column temperature were used to design mathematical models. Central composite design (CCD) was used to study the response surface methodology and to study in depth the effects of these independent factors. Desirability function was used to simultaneously optimize the retention time and resolution of SAXA and DAPA. The optimized and predicted data from contour diagram consisted of acetonitrile and ortho phosphoric acid (0.1%) in the ratio of 50:50 respectively, at a flow rate of 0.98 ml/min and column temperature 31.4 °C. Using these optimum conditions baseline separation of both drugs with good resolution and run time of less than 6 min were achieved. The optimized assay conditions were validated according to ICH guidelines. Hence, the results clearly showed that Quality by design approach could be successfully applied to optimize RP-HPLC method for simultaneous estimation of SAXA and DAPA.
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Préparations pharmaceutiques/classification , Chromatographie en phase liquide à haute performance/méthodes , Diabète de type 2/classification , Comprimés/administration et posologie , Formes posologiques , Optimisation du Processus/méthodesRÉSUMÉ
Objetivo: Avaliar a capacidade de remediação do DEET em meio líquido, pelo fungo de decomposição branca Pleurotus ostreatus usando como indutor enzimático os resíduos sólidos do cacau e realizar bioensaios de toxicidade com as amostras pós-tratamento, para aplicações em tratamentos de águas. Método: Foi realizada a produção enzimática com resíduos do Cacau. A biorremediação com o caldo enzimático foi realizada em erlenmeyers de 250mL, contendo a solução do composto, tampão acetato de sódio pH 5 e o caldo batata, incubados à 28°C, com rotação de 120 rpm, por 48 horas. Já com o fungo ativo, o mesmo foi incubado a 28 ºC e teve em seu meio a adição do composto. As amostras foram quantificados em Cromatografia líquida de alta performance (CLAE). O teste de adsorção foi feito com o fungo autoclavado e analisado após 14 dias. Resultado: O composto se apresentou possivelmente tóxico e a remediação mostrou uma tendência linear de degradação com o fungo de 39%. Conclusão: Pleurotus ostreatus é um candidato promissor para o tratamento de contimanações geradas por DEET.
Objective: We evaluated the remediation capacity of DEET in liquid medium by the white decomposition fungus Pleurotus ostreatus using the solid residues of cocoa as an enzymatic inducer and performed toxicity bioassays with the post-treatment samples, for water treatment applications. Method: Enzymatic production with cocoa residues was performed. Bioremediation with the enzyme broth was performed in a 250mL erlenmeyer flasks, containing the solution of the compound, sodium acetate buffer pH 5 and the potato broth, incubated at 28 °C, with rotation of 120 rpm, for 48 hours. With the active fungus, the same was incubated at 28 ºC and had in its culture medium the addition of the compound. The samples were quantified in high performance liquid chromatography (HPLC). The adsorption test was performed with the autoclaved fungus and analyzed after 14 days. Results: The compound was possibly toxic and the remediation showed a linear tendency of degradation of 39% with the fungus. Conclusion: Pleurotus ostreatus is a promising candidate for the treatment of contaminants generated by DEET.
Sujet(s)
Chromatographie en phase liquide à haute performanceRÉSUMÉ
Objective@#Study on the improvement of quality standard for Zicao ointment.@*Methods@#The content of shikonin and β,β’-dimethylacrylshikonin was determined by high performance liquid chromatogram (HPLC). The column was Inertsil ODS-3 C18 column (4.6 mm×250 mm, 5 μm). The mobile phase comprising of acetonitrile-0.05% formic acid solution (70:30). The flow rate was 1.0 ml/min, the detection wavelength was 275 nm, and the column temperature was 25 ℃.@*Results@#The linear range of shikonin and β,β’-dimethylacrylshikonin were 0.04-0.81 μg (r=0.999 5) and 0.41-10.36 μg (r=0.999 2). The average recovery were 103.32% and 101.80%, the RSD were 1.96% and 2.11%.@*Conclusions@#The method is simple, accurate and sensitive to control the quality of Zicao ointment.
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Objective To establish HPLC characteristic chromatograms method for quality control of Saponins in Panacis Majoris Rhizoma.Methods The HPLC method was conducted based on a Agilent Poroshell 120 C18 (4.6 mm × 100 mm, 2.7μm) column, with acetonitrile-0.1％ orthophosphoric acid as mobile phase by gradient elution. The flow rate was 1.0 ml/min. The detection wavelength was at 203 nm and the column temperature was 35℃. Results Eight samples from different sources were analyzed by HPLC. Fourteen main marker peaks were selected in the Characteristic spectrum and four peaks of saponins were identified. The similarity of seven samples was over 0.9.Conclusions The precision, repeatability and stability method was standardized, and this method can be used for quality control of Panacis Majoris Rhizoma.
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Objective To investigate the metabolomics characteristic of neuromyelities optica spectrum disorder (NMOSD) in plasma and cerebrospinal fluid.Methods Ultra high performance liquid chromatography-mass spectrometry (UHPLC-MS) was used to identify plasma metabolites in 16 patients with NMOSD and 8 healthy controls.At the same time,the identification of metabolites in cerebrospinal fluid of 8 NMOSD patients and 5 healthy controls was completed.Differential metabolites screening and metabolomic pathway analysis were performed by diversified data analysis methods.Results Compared with healthy control group,the content of 8 substances such as Cis.8.11.14.Eicosatrienoic acid in the plasma of NMOSD patients was increased.The content of 8 substances such as L-glutamine acid were decreased.There was no significant difference in the metabolites between Aquaporin 4 (AQP-4) antibody positive and negative NMOSD plasma.The content of 6 substances such as 3-hydroxybutyric acid in cerebrospinal fluid of patients with NMOSD was reduced.Conclusions The distribution of metabolites in plasma between NMOSD patients and healthy controls was significantly different.There was no significant difference in metabolites between AQP-4 antibody positive and negative NMOSD plasma.There are some differences in metabolites between cerebrospinal fluid of NMOSD patients and healthy controls.A variety of amino acid abnormalities,sphingomyelin dysfunction,energy metabolism and mitochondrial dysfunction were involved in the pathogenesis of NMOSD.
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Objective To establish a method for determining five isoquinoline alkaloids (including berberine,jatrorrhizine,jatrorrhizind,coptisine and palmatine) in Erhuang Xiaoyan tablets simultaneously.Methods Determination by reversed phase high performance liquid chromatography (RP-HPLC).Column:YMC-Pack ODS-AM (4.6 mm× 250.0 mm,5 μm);mobile phase:elution with 0.1％ (v/v) formic acid aqueous solution-acetonitrile mobile phase gradient;flow rate:0.4 mL/min;injection volume:10 μL;detection wavelength:345 nm;column temperature:27 ℃.Results Five of isoquinoline alkaloids in Erhuang Xiaoyan tablets were separated perfectly.The good linear relationships were obtained in the following ranges,1.526 4-5.342 4μg (r=0.999 9) for berberine,0.403 2-1.411 2 μg (r=0.999 6) for palmatine,0.309 7-1.083 9 μg (r=0.999 8) for coptisine,0.111 2-0.389 2 μg (r=0.999 8) for jatrorrhizine and 0.162 2-0.567 7 μg (r=0.999 6) for epiberberine.The recoveries were 98.69％,98.66％,98.67％,98.67％ and 98.67％,respectively.Conclusion The method possesses high feasibility,strong specificity,high accuracy and good reproducibility,which can be used for the quality control of Erhuang Xiaoyan tablets.
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Objective To study the qualitative and quantitative standard of Shengui anti-pruritic mixture. Mehtods Sophorae Flavescentis Radix, Cnidii Fructus, Polygoni Multiflori Radix, Astragali Radix, Paeoniae Radix Alba and Glycyrrhizae Radix Et Rhizoma were identified by TLC.The contents of matrine and ferulic acid were determined by HPLC. Results The TLC showed the obvious characteristics with the spots clear and well-separated with no interference by negative control. Matrine showed a good linear relationship at 0.062 5- 2.500 0 μg (r=0.999 9) with a good meeting of adding sample recovery test. Ferulic acid showed a good linear relationship at 0.021 0-0.840 0 μg (r=0.999 9) with a good meeting of adding sample recovery test. Conclusions The method is easy-operated and accurate which showed a good specificity for the quality control of Shengui anti-pruritic mixture.
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Objective To establish an HPLC method for simultaneous determination of Quercetin , Luteolin, pigenin in Matricaria Chamomila L.. Methods HPLC analysis was performed on Agilent Zorbox SB-C18 column (4.6 mm×250 mm, 5 μm) with the methanol and phosphoric acid as mobile phase in equal degree model, and the column temperature was set at 25 ℃, and the flow rate was 1.0 ml.min-1 and the detecting wave length was at 350 nm. Results The linear response ranges were from 0.25-4.04 μg of Quercetin (r=1.000, n=6) and from 0.25-3.98 μg of Luteolin (r=1.000, n=6) and 0.25-4.02 μg of Apigenin (r=1.000, n=6);and the recoveries, the precision and the stability RSD of Quercetin, Luteolin and Apigenin meet the requirements. The average recovery rate of quercetin, luteolin and apigenin was 93.64%, 95.85% and 95.40%, respectively. Conclusions All 3 samples in Matricaria Chamomila.L were determined by HPLC. The method is simple, rapid, and reproducible. It can be used as reference for the quality control of Matricaria Chamomila L..