高效液相色谱电生Mn(Ⅲ)化学发光法检测人体血清和尿样中的卡托普利

设计了一个HPLC在线电生Mn(Ⅲ)化学发光检测器, 实现在线电化学反应, 从而产生反应活性很高的初生态氧化剂Mn(Ⅲ), 并与色谱柱后CP混合产生化学发光. 同时还能够根据需要调节电极反应和发光反应两者的介质, 满足柱后发光反应的最佳环境. 在优化流动相和化学发光检测条件的基础上, 将该检测器应用于人体血清和尿液中CP的测定.     

Assay of free captopril in human plasma as monobromobimane derivative,using RPLC/(+)ESI/MS/MS: validation aspects and bioequivalence evaluation

A sensitive method for determination of free captopril as monobromobimane derivative in plasma samples is discussed. The internal standard (IS) was 5‐methoxy‐1H‐benzimidazole‐2‐thiol. Derivatization with monobromobimane immediately after blood collection and plasma preparation prevents oxidation of captopril to the corresponding disulfide compound and enhances the ionization yield. Consequently, derivatization enhances sample stability and detection sensitivity. Addition of the internal standard was made immediately after plasma preparation. The internal standard was also derivatized by monobromobimane, as it contains a thiol functional group. Preparation of plasma samples containing captopril and IS derivatives was based upon protein precipitation through addition of acetonitrile, in a volumetric ratio 1:2. The reversed‐phase liquid chromatographic separation was achieved on a rapid resolution cartridge Zorbax SB‐C₁₈, monitored through positive electrospray ionization and tandem MS detection using the multiple‐reaction monitoring mode. Transitions were 408–362 amu for the captopril derivative and 371–260 amu for the internal standard derivative. The kinetics of captopril oxidation to the corresponding disulfide compound in plasma matrix was also studied using the proposed method. A linear log–log calibration was obtained over the concentration interval 2.5–750 ng/mL. A low limit of quantitation in the 2.5 ng/mL range was obtained. The analytical method was fully validated and successfully applied in a three‐way, three‐period, single‐dose (50 mg), block‐randomized bioequivalence study for two pharmaceutical formulations (captopril LPH 25 and 50 mg) against the comparator Capoten 50 mg. Copyright © 2009 John Wiley & Sons, Ltd.     

Cationic cyanine as a near-infrared fluorescent probe for the determination of nucleic acids

A new method with a cationic near-IR cyanine as fluorescent probe was developed for the determination of nucleic acids. The near-IR cyanine shows maximum excitation and emission wavelengths at 765 and 790 nm, respectively, in aqueous solution. The method is based on the fluorescence decrease of near-IR cyanine in the presence of nucleic acids. Under optimal conditions, the ratio of fluorescence intensity in the absence and presence of nucleic acids was proportional to the concentration of nucleic acids over the range 0.10-1.2 microg/mL for CT (calf thymus) DNA or SM (salmon sperm) DNA, and 0.10-1.6 microg/mL for yeast RNA. The detection limits were 30 ng/mL for CT DNA, 25 ng/mL for SM DNA and 70 ng/mL for yeast RNA. The relative standard deviation (n = 6) was 2.1% for 500 ng/mL CT DNA, 2.4% for 500 ng/mL SM DNA and 2.7% for 500 ng/mL yeast RNA, respectively.     

Ionic liquid as a mobile phase additive in high‐performance liquid chromatography for the simultaneous determination of eleven fluorescent whitening agents in paper materials

In the present study, 11 4,4′‐diaminostilbene‐2,2′‐disulfonic acid based fluorescent whitening agents with different numbers of sulfonic acid groups were separated by using an ionic liquid as a mobile phase additive in high‐performance liquid chromatography with fluorescence detection. The effects of ionic liquid concentration, pH of mobile phase B, and composition of mobile phase A on the separation of fluorescent whitening agents were systematically investigated. The ionic liquid tetrabutylammonium tetrafluoroborate is superior to tetrabutylammomnium bromide for the separation of the fluorescent whitening agents. The optimal separation conditions were an ionic liquid concentration at 8 mM and the pH of mobile phase B at 8.5 with methanol as mobile phase A. The established method exhibited low limits of detection (0.04–0.07 ng/mL) and wide linearity ranges (0.30–20 ng/mL) with high linear correlation coefficients from 0.9994 to 0.9998. The optimized procedure was applied to analyze target analytes in paper samples with satisfactory results. Eleven target analytes were quantified, and the recoveries of spiked paper samples were in the range of 85–105% with the relative standard deviations from 2.1 to 5.1%. The obtained results indicated that the method was efficient for detection of 11 fluorescent whitening agents.     

基于BHHCT-Eu3+@SiO2荧光稀土二氧化硅纳米颗粒的免疫层析试纸条检测卡那霉素

采用微波辅助合成的荧光稀土二氧化硅纳米颗粒(BHHCT-Eu3+@SiO2)为标记物,建立了快速定量检测卡那霉素(Kana)残留的荧光免疫层析方法.实验结果表明,微波辅助合成的BHHCT-Eu3+@SiO2纳米颗粒呈球形,粒径约36 nm,具有良好的荧光发射性能,最大吸收波长和最大发射波长分别为343和615 nm.将BHHCT-Eu3+@SiO2与卡那霉素抗体(Kana-ab)通过醛基化葡聚糖交联,合成了荧光标记抗体Eu3+-Kana-ab,结合定量侧向层析读数仪,建立了牛奶中Kana残留的快速定量检测方法,对Kana的检出限(IC10)为0.85 ng/mL,半数抑制浓度(IC50)为12.76 ng/mL,检测范围(IC20-IC80)为3.0~76.0 ng/mL,牛奶中的Kana的加标回收率范围为93.7%~97.4%,RSD为3.1%~4.6%,与Kana类似物的交叉反应均<1%.牛奶中Kana残留的测定结果与ELISA方法相关性良好.     

Cationic cyanine as a near-infrared fluorescent probe for the determination of nucleic acids

A new method with a cationic near-IR cyanine as fluorescent probe was developed for the determination of nucleic acids. The near-IR cyanine shows maximum excitation and emission wavelengths at 765 and 790 nm, respectively, in aqueous solution. The method is based on the fluorescence decrease of near-IR cyanine in the presence of nucleic acids. Under optimal conditions, the ratio of fluorescence intensity in the absence and presence of nucleic acids was proportional to the concentration of ¶nucleic acids over the range 0.10–1.2 μg/mL for CT (calf thymus) DNA or SM (salmon sperm) DNA, and 0.10–¶1.6 μg/mL for yeast RNA. The detection limits were ¶30 ng/mL for CT DNA, 25 ng/mL for SM DNA and ¶70 ng/mL for yeast RNA. The relative standard deviation (n = 6) was 2.1% for 500 ng/mL CT DNA, 2.4% for ¶500 ng/mL SM DNA and 2.7% for 500 ng/mL yeast RNA, respectively.     

Highly sensitive determination and validation of gabapentin in pharmaceutical preparations by HPLC with 4-fluoro-7-nitrobenzofurazan derivatization and fluorescence detection

A sensitive HPLC method with pre-column fluorescence derivatization using 4-Fluoro-7-Nitrobenzofurazan (NBD-F) has been developed for the determination of gabapentin in pharmaceutical preparations. The method is based on the derivatization of gabapentin with (NBD-F) in borate buffer of pH 9.5 to yield a yellow, fluorescent product. The HPLC separation was achieved on a Inertsil C(18) column (250 mm × 4.6 mm) using a mobile phase of methanol water (80:20, v/v) solvent system at 1.2 mL/min flow rate. Mexiletine was used as the internal standard. The fluorometric detector was operated at 458 nm (excitation) and 521 nm (emission). The assay was linear over the concentration range of 5 50 ng/mL. The method was validated for specificity, linearity, limit of detection, limit of quantification, precision, accuracy, robustness. Moreover, the method was found to be sensitive with a low limit of detection (0.85 ng/mL) and limit of quantitation (2.55 ng/mL). The results of the developed procedure for gabapentin content in capsules were compared with those by the official method (USP 32). Statistical analysis by t- and F-tests, showed no significant difference at 95 confidence level between the two proposed methods.     

Synthesis of 1,5-bis(4,6-dichloro-1,3,5-triazinylamino)naphthalene and its application to spectrofluorimetric determination of aniline

A new fluorescent reagent 1,5-bis(4,6-dichloro-1,3,5-triazinylamino)naphthalene (DTAN) was synthesized. The optimum conditions of fluorescent reaction of this reagent with aniline (PA) were also investigated. Based on this reaction, a new spectrofluorimetric method was developed for the determination of aniline. The fluorescent intensity was directly proportional to the concentration of aniline in the ranges 0.05-2.0 microg mL(-1) and 2.0-50 microg mL(-1) with the detection limits of 34 ng mL(-1) and 90 ng mL(-1). This method is simple, practical and can afford good precision and accuracy and can be successfully applied to assess aniline in water samples. A possible mechanism of the change of fluorescence intensity introduced by putting the aniline into the system is also discussed.     

Development of Liposome-Based Immunoassay for the Detection of Cardiac Troponin I

Cardiovascular diseases (CVDs) are one of the foremost causes of mortality in intensive care units worldwide. The development of a rapid method to quantify cardiac troponin I (cTnI)—the gold-standard biomarker of myocardial infarction (MI) (or “heart attack”)—becomes crucial in the early diagnosis and treatment of myocardial infarction (MI). This study investigates the development of an efficient fluorescent “sandwich” immunoassay using liposome-based fluorescent signal amplification and thereby enables the sensing and quantification of serum-cTnI at a concentration relevant to clinical settings. The calcein-loaded liposomes were utilized as fluorescent nano vehicles, and these have exhibited appropriate stability and efficient fluorescent properties. The standardized assay was sensitive and selective towards cTnI in both physiological buffer solutions and spiked human serum samples. The novel assay presented noble analytical results with sound dynamic linearity over a wide concentration range of 0 to 320 ng/mL and a detection limit of 6.5 ng/mL for cTnI in the spiked human serum.     

Nonaqueous capillary electrophoresis coupled with laser-induced native fluorescence detection for the analysis of berberine, palmatine, and jatrorrhizine in Chinese herbal medicines

LIF detection is one of the most sensitive detection methods for CE. However, its application is limited because the analyte is usually required to be derivatized with a fluorescent label. As a result, LIF is seldom used to analyze active ingredients in plants. In this work, we introduce a rapid, simple, and sensitive method of nonaqueous CE (NACE) coupled with laser-induced native fluorescence detection for the simultaneous analysis of berberine, palmatine, and jatrorrhizine. This method skillfully utilizes the native fluorescence of these alkaloids and requires no troublesome fluorescent derivatization. As these alkaloids can fluoresce to some degree, they were simply detected by a commercially available 488 nm Ar+ laser. The native fluorescence of the analytes was greatly enhanced by nonaqueous media. Compared with the reported UV detection method, much lower LOD was achieved (6.0 ng/mL for berberine, 7.5 ng/mL for palmatine, and 380 ng/mL for jatrorrhizine). This method was successfully applied to analyze berberine, palmatine, and jatrorrhizine in two Chinese herbal medicines, Rhizoma coptidis and Caulis mahoniae.     

硫胺素-亚硝酸根体系的荧光性质研究及应用

研究了在碱性溶液中硫胺素（VB1）与亚硝酸根的荧光反应,建立了荧光光度法测定硫胺素的新方法。方法线性范围为1．28－324ng/mL,检出限0．52ng/mL,对15个浓度为100ng/mL的硫胺素溶液进行测定,相对标准偏差为1．4％。应用于药物中硫胺素的测定,回收率在96．5％－101．8％之间。     

Determination of bupropion using liquid chromatography with fluorescence detection in pharmaceutical preparations, human plasma and human urine

A novel pre-column derivatization reversed-phase high-performance liquid chromatography with fluorescence detection is described for the determination of bupropion in pharmaceutical preparation, human plasma and human urine using mexiletine as internal standard. The proposed method is based on the reaction of 4-chloro-7-nitrobenzofurazan (NBD-Cl) with bupropion to produce a fluorescent derivative. The derivative formed is monitored on a C18 (150 mm × 4.6 mm i.d., 5 μm) column using a mobile phase consisting of methanol-water 75:25 (v/v), at a flow-rate of 1.2 mL/min and detected fluorimetrically at λ(ex) = 458 and λ(em) = 533 nm. The assay was linear over the concentration ranges of 5-500 and 10-500 ng/mL for plasma and urine, respectively. The limits of detection and quantification were calculated to be 0.24 and 0.72 ng/mL for plasma and urine, respectively (inter-day results). The recoveries obtained for plasma and urine were 97.12% ± 0.45 and 96.00% ± 0.45, respectively. The method presents good performance in terms of precision, accuracy, specificity, linearity, detection and quantification limits and robustness. The proposed method is applied to determine bupropion in commercially available tablets. The results were compared with an ultraviolet spectrophotometry method using t- and F-tests.     

Long‐chain ionic liquid based mixed hemimicelles and magnetic dispersed solid‐phase extraction for the extraction of fluorescent whitening agents in paper materials

A novel mixed hemimicelles and magnetic dispersive solid‐phase extraction method based on long‐chain ionic liquids for the extraction of five fluorescent whitening agents was established. The factors influenced on extraction efficiency were investigated. Under the optimal conditions, namely, the pH of sample solution at 8.0, the concentration of long chain ionic liquid at 0.5 mmol/L, the amount of Fe₃O₄ nanoparticle at 12 mg, extraction time at 10 min, pH 6.0 of methanol as eluent, and the desorption time at 1 min, satisfactory results were obtained. Wide linear ranges (0.02–10 ng/mL) and good linearity were attained (0.9997–0.9999). The intraday and interday RSDs were 2.1–8.3%. Limits of detection were 0.004–0.01 ng/mL, which were decreased by almost an order of magnitude compared to direct detection without extraction. The present method was applied to extract the fluorescent whitening agents in two kinds of paper samples, obtaining satisfactory results. All showed results illustrated that the detection sensitivity was improved and the proposed method was a good choice for the enriching and monitoring of trace fluorescent whitening agents.     

Application of potassium ferricyanide in the spectrophotometric determination of captopril

A novel method for the determination of captopril by speetrophotometer is described in this paper. The experiment is based on the fact that Fe(Ⅲ) is reduced to Fe(Ⅱ) by captopril, then the in situ formed Fe(Ⅱ) reacts with potassium ferricyanide to give the soluble prussian blue at pH 4.00, and its maximal adsorption wavelength (λmax) is 735 nm. Good linear relationship is obtained between the absorbance and the concentration of captopril in the wide range of 0.05-20 μg/mL. The linear regression equation is A = -0.04314 + 0.11423C (μg/mL) with a correlation coefficient R = 0.9998. The detection limit (3a/k) is 0.04 μg/mL, the molar absorption coefficient is 2.5×104 L/mol cm. By mensurating the absorbance of soluble prussian blue, the indirect determination of eaptopril can be obtained. This method has been successfully applied to determination of captopril in pharmaceutical samples.Analytical results obtained are satisfactory.     

Spectrofluorimetric Determination and Validation of Biotin in Pure and Dosage Form via Derivatization with 4-Fluoro-7-nitrobenzofurazan

A simple and highly sensitive spectrofluorimetric method was developed for the determination of biotin in pure and dosage form. The method is based on the derivatization of biotin with 4‐fluoro‐7‐nitrobenzofurazan in borate buffer of pH 9.0 to yield a yellow, fluorescent product. The various chemical conditions that affected the reaction were studied. The method was validated for specificity, linearity, precision, accuracy, recovery and robustness. At optimized experimental conditions, a linear relationship between the fluorescence intensity of the concentration of biotin is observed in the range 45–450 ng/mL. Limit of detection and quantification were 0.038 and 0.114 ng/mL, respectively. The percentage mean recovery was 99.96. The proposed procedure was successfully applied to the determination of biotin in its dosage form with mean recovery of 101.23±1.22 for biotin tablets. The results obtained were in good agreement with those obtained by the reference method.     

Improved method for determination of abamectin and ivermectin in cattle plasma.

A liquid chromatographic (LC) method was developed for determination of abamectin (ABM) and ivermectin (IVM) in cattle plasma. The sample was extracted with acetonitrile and cleaned up on an alumina column. After conversion to stable fluorescent derivative with trifluoroacetic anhydride and N-methylimidazole, the sample was analyzed by LC with fluorescence detection (Ex 365 nm and Em 475 nm). Doramectin was used as an internal standard. Recoveries ranged from 91.2 to 100.7% for IVM and from 87.0 to 98.7% for ABM, with 1-50 ng/mL fortified samples. The coefficients of variation were <10.1%. The limit of detection was 0.02 ng/mL for ABM and IVM in 1.0 mL samples.     

Separation and determination of closely related lantibiotics by micellar electrokinetic chromatography

A sensitive micellar electrokinetic chromatography (MEKC) method was developed for the separation and determination of four closely related lantibiotics: gallidermin, cinnamycin, duramycin and nisin. Factors affecting the separation of the lantibiotics such as pH, phosphate buffer concentration, SDS concentration and wavelength for UV detection were investigated. By optimizing these experimental conditions, successful separation was achieved between class 1A lantibiotics (nisin and gallidermin) and class 1B lantibiotics (duramycin and cinnamycin). The four lantibiotics were separated within 12 min in 50 mM phosphate buffer at pH 3.95 ± 0.1 containing 80 mM SDS with UV detection of 214 nm. The LOD (S/N = 3) were 61 ng/mL for gallidermin, 57 ng/mL for cinnamycin, 55 ng/mL for duramycin and 58 ng/mL for nisin. The method was successfully applied to real samples such as fermentation broth, bovine colostrum and predrop beer. This method yielded satisfactory results, with quantitative recoveries of spiked lantibiotics in the three samples ranging from 86.1 to 99.6%.     

The Application of Gibberellic Acid to the Determination of Trace Amounts of Lead by Spectrofluorimetry

The ionization constant of fluorescent reagent gibberellic acid (GA) was established spectrophotometrically. The fluorescent reaction of this reagent with lead was studied. Based on this chelation reaction, a sensitive, direct spectrofluorimetric method for the determination of trace lead with use of GA has been developed. The reaction conditions for the fluorescence system of lead with GA were studied. The lead ion can form a stable binary chelate with GA, having a ratio of 1:2 in the pH range 7.0‐8.0. The maximum excitation and emission wavelengths are 205.0 nm and 308.8 nm for the lead chelate, respectively. The reaction is instantaneous and the fluorescence intensity of the lead chelate remains stable from 20 to 150 min. Under the optimal experimental conditions the fluorescence intensity is a linear function of concentration in the range 1.0‐10.0 ng/mL of lead and the detection limit is 0.52 ng/mL of lead. Interferences of other ions were studied. The method has been successfully applied to the determination of lead in common paint.     

A kinetic fluorometric method for the determination of nucleic acids using a ternary equilibrium system of nucleic acids—iron (III) tetracarboxy phthalocyanine-poly-lysine coupled with the oxidation reaction between hydrogen peroxide and dl-tyrosine

Using the oxidation reaction between hydrogen peroxide and dl-tyrosine as fluorescence indication, the evident tuning effect of nucleic acids on catalytic activity of mimetic enzyme iron (III) tetracarboxy phthalocyanine (FeC₄Pc) in the presence of poly-lysine was observed and studied. The oxidation reaction between hydrogen peroxide and dl-tyrosine with FeC₄Pc as catalyst gave an intensively fluorescent compound, which has an excitation wavelength of 325 nm and an emission wavelength of 418 nm. The fluorescence was quenched by a proper concentration of poly-lysine due to its association with FeC₄Pc and consequently the descent of the catalytic activity of FeC₄Pc, but recovered by addition of nucleic acids. Under optimal conditions, the recovered fluorescence is proportional to the concentration of nucleic acids. Based on the fact, a kinetic fluorescent method was developed for the determination of nucleic acids. The calibration graphs are linear over the range 10-2000 ng/mL both for fish sperm DNA (FS DNA) and calf thymus DNA (CT DNA). The corresponding detection limits are 1.04 ng/mL for FS DNA and 1.18 ng/mL for CT DNA, respectively. Four synthetic and three real nucleic acid samples were determined with satisfactory results.     

High-sensitivity analysis of cyanide by capillary electrophoresis with fluorescence detection.

A capillary electrophoretic method for a high-sensitivity analysis of cyanide has been developed. Cyanide was derivatized with 2,3-naphthalenedialdehyde and taurine to give a fluorescent product of 1-cyanobenz[f]isoindole. This compound was detected with high sensitivity by fluorescence detection. The detection limit was 0.1 ng/mL, and the calibration curve was linear over the range 0.1-200 ng/mL. The precision of the migration time of within-run assays (n = 6) of 1 ng/mL cyanide standard solution was 0.14%. The precision of the peak area for the same runs was 1.0%. This method was applicable to blood analysis. Detection of the cyanide derivative by UV was also examined.