Fluorescent Reaction Research Articles

Polony DNA Sequencing

Polony DNA sequencing provides an inexpensive, accurate, high-throughput way to resequence genomes of interest by comparison to a reference genome. Mate-paired in vitro shotgun genomic libraries are produced and clonally amplified on microbeads by emulsion PCR. These serve as templates for sequencing by fluorescent nonamer ligation reactions on a microscope slide. Each sequencing run results in millions of 26-bp reads that can be aligned to the reference genome, allowing the identification of differences between sequences.

Front face fluorescence spectroscopy as a tool for the assessment of egg freshness during storage at a temperature of 12.2 °C and 87% relative humidity

The objective of this study was to investigate intrinsic fluorophores of thick albumen and egg yolk in order to assess egg freshness during storage at a temperature of 12.2 °C and 87% relative humidity (RH). A total of 126 intact brown-shelled eggs of the same flock (29 weeks of age) were stored for 1, 6, 8, 12, 15, 20, 22, 26, 29, 33, 40, 47 and 55 days. The emission fluorescence spectra of aromatic amino acids and nucleic acids (AAA + NA) (excitation: 250 nm; emission: 280–450 nm), fluorescent Maillard reaction products (FMRP) (excitation: 360 nm; emission: 380–580 nm) and the excitation spectra of vitamin A (emission: 410 nm; excitation: 270–350 nm) were scanned on thick albumen and egg yolk. Among the intrinsic fluorophores, only the principal component analysis (PCA) applied on the vitamin A fluorescence spectra allowed a good identification of eggs as a function of their storage time. Factorial discriminant analysis (FDA) was then applied on the first five principal components (PCs) of the PCA applied on each spectral data set. Regarding AAA + NA recorded on thick albumen, correct classification of 69.4% and 63.9% was observed for the calibration and validation sets, respectively. Quite similar results were obtained on AAA + NA scanned on egg yolks. The best results were obtained with vitamin A fluorescence spectra since 97.7% and 85.7% of the calibration and validation sets was obtained, respectively. These results showed that vitamin A fluorescence spectra provide useful fingerprints, mainly allow the identification of eggs during storage and could be considered as a powerful intrinsic probe for the evaluation of egg freshness.

Highly Sensitive Spectrofluorimetric Determination of Ephedrine Hydrochloride in Pharmaceutical Preparations

A sensitive and specific spectrofluorimetry method was developed and validated for the quantification of ephedrine (EP) in pharmaceutical preparations. The method is based on the fluorescent enhancing reaction of EP with 7-chloro-4-nitrobenzofurazan (NBD-C1; derivatization reagent), in borate buffer of pH 9 to yield a yellow, fluorescent product. Under these experimental conditions, the derivatized product of EP had excitation and emission wavelength maxima at 458 and 516 nm, respectively. The linear range of this method was 20-2500 ng/mL. The detection limit was 7.3 ng/mL EP. Intra- and interday precisions of the assay at 3 concentrations within this range were 0.037-1.77%. The low relative standard deviation values indicate good precision, and high recovery values indicate excellent accuracy of the method. The proposed method was applied to the determination of the examined drugs in pharmaceutical formulations, and the results indicate that the method is equally as accurate, precise, and reproducible as the official method.

Relative quantification of cytokeratin 19 transcription in oral squamous cell carcinoma tissues by fluorescent quantitive real-time RT-PCR

To investigate the relative quantification of cytokeratin 19 transcription in oral squamous cell carcinoma tissues by fluorescent quantitive real-time reverse transcription-polymerase chain reaction ((RT-PCR). CK19mRNA level was detected by fluorescent quantitive real-time RT-PCR in cancerous and para-cancerous tissues from 31 oral squamous cell carcinoma patients. According to the 2(-DeltaDeltaCt) equation, the relative quantification fold of CK19mRNA level was calculated in cancerous tissues compared with para-cancerous tissues. CK19mRNA levels in cancerous tissues were 2.21 folds higher than those in para-cancerous tissues, and the amplicon was specific. CK19mRNA level in cancerous tissue correlated significantly with pathological differentiation degree, the poorer the differentiation was, the higher the CK19mRNA level became. Fluorescent quantitive real-time RT-PCR is accurate and reliable in the detection of relative quantification of CK19 transcription in oral squamous cell carcinoma tissues.

High Sensitive Fluorophotometric Determination of Nucleic Acids with Pyronine G Sensitized by N,N‐Dimethylformamide

In Tris-HCl buffer (pH 8.0), the fluorescence of Pyronine G emitted at 552 nm was quenched by nucleic acids when excited at 525 nm. Adding N, N-dimethylformamide (DMF) as a sensitive media can enhance the sensitivity greatly. Based on the fluorescence reactions sensitive fluorimetric methods for nucleic acids at nanogram levels were proposed. Under the optimum conditions, the calibration curves were linear in the range of 0.0032 - 2.5 microg mL(-1) for ct DNA and 0.0024 - 2.5 microg mL(-1) for hs DNA. The limits of determination were 3.2 ng mL(-1) and 2.4 ng mL(-1) respectively. This method has good selectivity and high sensitivity. It has been applied to the determination of DNA in the synthetic samples and real samples with satisfactory results.

Functional modulation of human intestinal epithelial cell responses by Bifidobacterium infantis and Lactobacillus salivarius

Intestinal epithelial cells (IECs) and dendritic cells (DCs) play a pivotal role in antigen sampling and the maintenance of gut homeostasis. However, the interaction of commensal bacteria with the intestinal surface remains incompletely understood. Here we investigated immune cell responses to commensal and pathogenic bacteria. HT-29 human IECs were incubated with Bifidobacterium infantis 35624, Lactobacillus salivarius UCC118 or Salmonella typhimurium UK1 for varying times, or were pretreated with a probiotic for 2 hr prior to stimulation with S. typhimurium or flagellin. Gene arrays were used to examine inflammatory gene expression. Nuclear factor (NF)-kappaB activation, interleukin (IL)-8 secretion, pathogen adherence to IECs, and mucin-3 (MUC3) and E-cadherin gene expression were assayed by TransAM assay, enzyme-linked immunosorbent assay (ELISA), fluorescence, and real-time reverse transcriptase-polymerase chain reaction (RT-PCR), respectively. IL-10 and tumour necrosis factor (TNF)-alpha secretion by bacteria-treated peripheral blood-derived DCs were measured using ELISA. S. typhimurium increased expression of 36 of the 847 immune-related genes assayed, including NF-kappaB and IL-8. The commensal bacteria did not alter expression levels of any of the 847 genes. However, B. infantis and L. salivarius attenuated both IL-8 secretion at baseline and S. typhimurium-induced pro-inflammatory responses. B. infantis also limited flagellin-induced IL-8 protein secretion. The commensal bacteria did not increase MUC3or E-cadherin expression, or interfere with pathogen binding to HT-29 cells, but they did stimulate IL-10 and TNF-alpha secretion by DCs. The data demonstrate that, although the intestinal epithelium is immunologically quiescent when it encounters B. infantis or L. salivarius, these commensal bacteria exert immunomodulatory effects on intestinal immune cells that mediate host responses to flagellin and enteric pathogens.

Spectrofluorimetric determination of trace amounts of cadmium with 2,4-dihydroxyacetophenone benzoylhydrazone

A new fluorescent reagent 2,4-dihydroxyacetophenone benzoylhydrazone (DHAPBH) was synthesized and its ionization constants were established spectrophotometrically. The fluorescent reaction of this reagent with cadmium was studied. The cadmium ion can form a stable binary chelate with DHAPBH, having a mole ratio of 1:2 in the pH range of 3.6–4.4. Based on this chelation reaction, a highly sensitive spectrofluorimetric method was developed for determination of trace amounts of cadmium in a water-ethanol (7:3 in volume ratio) medium at pH = 4.0. Under these conditions, the Cd-DHAPBH complex has excitation and emission maxima at 306 and 437 nm, respectively. The linear range of the method is from 0.36 to 140 μg·L −1 and the detection limit is 0.27 μg·L −1 of cadmium. Interferences of other ions were studied. The method was successfully applied to the determination of cadmium in mineral water, tap water, river water, and wastewater.

Highly Sensitive Spectrofluorometric Determination of Trace Amounts of Mercury with a New Fluorescent Reagent, 2-Hydroxy-1-naphthaldehydene-8-aminoquinoline

A new fluorescent reagent, 2-hydroxy-1-naphthaldehydene-8-aminoquinoline (HNAAQ), was synthesized. The fluorescent reaction of this reagent with mercury was also studied. Based on this chelation, a highly sensitive spectrofluorometric method was developed for the determination of trace amounts of mercury in a water-ethanol (5 + 1, v/v) medium at pH 8.0. Under these conditions, the Hg-HNAAQ complex has excitation and emission maxima at 406 and 445 nm, respectively. The linear range of the method is from 0 to 16 microg L(-1) and the detection limit is 0.056 microg L(-1) of mercury. The interference of other ions was studied. In order to enhance the selectivity in the determination of mercury by the present method, we also applied the separation of mercury by distillation. Thus, the selectivity of the method could be increased remarkably. The procedure can be easily performed, and affords good precision and accuracy. This method has been successfully applied to the determination of mercury in waste water and prawns.

The application of glyoxal-bis-(2-hydroxyanil) to the determination of trace amounts of cadmium by spectrofluorimetry

The fluorescent reaction of the reagent glyoxal-bis-(2-hydroxyanil) (GBHA) with cadmium was studied. Based on this chelation reaction, a sensitive, rapid, direct spectrofluorimetric method for the determination of trace cadmium with GBHA has been developed. The reaction conditions for the fluorescence system of cadmium with GBHA were established. The cadmium ion can form a stable chelate with GBHA, having a ratio of 1:3 in the 12.0 -13.0 pH range. The maximum excitation and emission wavelengths are 227.4 nm and 320.0 nm for the cadmium chelate, respectively. The fluorescence intensity of the cadmium 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-1 of cadmium and the detection limit is 0.65 ng mL-1 of cadmium. Interferences of foreign ions were studied. The method has been successfully applied to the determination of cadmium in the sewage.

A new terbium(III) chelate as an efficient singlet oxygen fluorescence probe

A terbium(III) chelate fluorescence probe for detection of singlet oxygen ( 1O 2) in aqueous media, N,N,N 1 ,N 1 -[2,6-bis(3′-aminomethyl-1′-pyrazolyl)-4-(9″-anthryl)pyridine] tetrakis (acetate)–Tb 3+ (PATA-Tb 3+), was designed and synthesized. The new chelate is highly water soluble, is almost nonfluorescent, and can specifically react with 1O 2 to yield a strongly fluorescent chelate, the endoperoxide of PATA-Tb 3+, accompanied by a remarkable increase in the fluorescence quantum yield from 0.46 to 10.5%. The long fluorescence lifetime of the endoperoxide (2.76 ms) allows the probe to be used favorably for time-resolved fluorescence detection of 1O 2. The studies of fluorescence property and reaction specificity indicate that the new probe is highly sensitive and selective for 1O 2. The probe was used for quantitative detection of 1O 2 generated from a MoO 4 2−–H 2O 2 system to give a detection limit of 10.8 nM. In addition, the good applicability of the probe was demonstrated by the real-time monitoring of the kinetic process of 1O 2 generation in a horseradish peroxidase-catalyzed oxidation system of indole-3-acetic acid in a weakly acidic buffer.

Development of a rapid method based on front face fluorescence spectroscopy for the monitoring of egg freshness: 1—evolution of thick and thin egg albumens

C � Springer-Verlag 2005 Abstract In this preliminary study, the intrinsic fluores- cence of thick and thin egg albumens was evaluated as a possible rapid method for the monitoring of egg freshness. The fluorescence emission spectra of tryptophan residues (excitation: 290 nm; emission: 305-430 nm) of proteins and fluorescent Maillard reaction products (excitation: 360; emission: 380-580 nm) were recorded directly on thick and thin albumen samples within 2-3, 4, 5, 9, 10, 12, 16, 18, 23, 25 and 29 days of storage. Principal component analy- sis (PCA) and factorial discriminant analysis (FDA) were applied to the spectra data sets. Considering tryptophan fluorescence spectra recorded on thick egg albumen, cor- rect classification was observed for 62.8 and 54.3% for the calibration and the validation sets, respectively. Better clas- sification was obtained from thin egg albumen since 67.3 and 69.1% of samples were correctly classified. Consid- ering fluorescent Maillard reaction products, the similarity map determined by the principal components (PCs) 1 and 2 showed a discrimination of eggs as a function of their storage time on both thick and thin albumens. The percent- age of samples correctly classified into four groups by the FDA was 97.4 and 91.4% for the calibration and validation thick albumen samples, respectively. It was concluded that fluorescent Maillard reaction products could be considered as fingerprints that may allow the discrimination between fresh and aged eggs.

Spectrofluorimetric Determination of Drugs Containing Active Methylene Group Using N1-Methyl Nicotinamide Chloride as a Fluorigenic Agent

A spectrofluorimetric method was described for the determination of drugs containing active methylene groups adjacent to carbonyl groups. The method was applied successfully to the determination of three life saving cardiovascular drugs, with narrow therapeutic indices: pentoxifylline (I), propafenone hydrochloride (II) and acebutolol hydrochloride (III), in laboratory-prepared mixtures, in commercial tablets and in plasma samples. The method involved the reaction of each of the tested drugs with N1-methyl nicotinamide chloride (NMNCl) in the presence of alkali, followed by addition of formic acid, where highly fluorescent reaction products were produced. The produced fluorescence were measured quantitatively at 472 nm (lambdaex 352 nm), 409 nm (lambdaex 310 nm) and 451 nm (lambdaex 266 nm) for (I), (II), and (III) respectively. The method was linear over concentration ranges of 10-1000 microg/ml , 0.2-12 microg/ml and 0.08-10 microg/ml in standard solutions for (I), (II), and (III) respectively. In spiked human plasma samples, calibration graphs were linear over concentration ranges of 20-1000 microg/ml, 0.2-15 microg/ml and 0.08-10 microg/ml for (I), (II), and (III) respectively. The method showed good accuracy, specificity and precision in both laboratory-prepared mixtures and spiked human plasma samples. The proposed method is simple, with low instrumentation requirements, suitable for quality control application, bioavailability and bioequivalency studies.

Observation of circulating tumour cells in patients with non-small cell lung cancer by real-time fluorescent quantitative reverse transcriptase-polymerase chain reaction in peroperative period

To assess whether surgical manoeuvre or resection of lung cancer could lead to haematogenous dissemination of malignant cells. In the mean time, the relationship between the sequence of vessel ligation and the haematogenous dissemination of cancer cells during operation was determined. Exploiting cytokeratin 19 (CK19)/carcinoembryonic antigen (CEA) mRNA as markers, 69 peripheral blood samples were collected from 23 consecutive patients with non-small cell lung cancer (NSCLC) who underwent surgical resection with curative intention in preoperative, intraoperative and postoperative period, respectively. Before the operation, all patients were randomly assigned to one of the two surgical procedure groups according to the order of vessel ligation, PV-first group and PA-first group. Additionally, the ten patients with benign lung disease served as control subjects undergoing surgical resection. The quantity and timing of the shedding of lung cancer cells into the circulation of patients were also monitored by fluorescent quantitative-reverse transcriptase-polymerase chain reaction before, during and after surgery. (1) The CK19 diagnostic test: the value of CK19 mRNA in operation was significantly higher than that of preoperation (5.246+/-0.196 vs. 4.472+/-0.164, P=0.000) and postoperation (5.246+/-0.196 vs. 4.694+/-0.177, P=0.013). The values between adenocarcinoma and squamous carcinoma were strikingly different (4.9110+/-1.0315 vs. 4.1891+/-0.4126, t=2.364, P=0.028). The values between PV-first group and PA-first group during perioperative period appear to be different (4.503 vs. 5.085, P=0.086). Before operation, of the 23 cases studied, 14 cases were positive (60.9%). Surprisingly, circulating epithelial cells were detected in two patients resected for benign lung disease. (2) The CEA diagnostic test: the level of CEA mRNA ascended continuously within this period. The postoperative values were significantly higher than those of preoperation (4.874 vs. 4.483, P=0.000) and those of operative day (4.874 vs. 4.537, P=0.000). The values between PV-first group and PA-first group appear to reach statistical significance (4.397 vs. 4.817, P=0.075). At the same time, there was a correlation between preoperative T-stage and perioperative CEA mRNA (4.267 vs. 4.760, P=0.025). Among the 23 cases, 10 cases were positive (43.5%). Both patients with benign lung disease served as control subjects undergoing surgical resection and the volunteers were negative. A considerable proportion of patients who appear to have resectable NSCLC might be regarded as having systemic disease, which is often undetectable by current tumour staging method. In terms of a marker used for the NSCLC patients who undergo operation, CEA is more suitable than CK19. The CK19-expressing epithelial cells are released intraoperatively into the circulation, meanwhile CEA-expressing tumour cells are disseminated mostly postoperatively. Surgical manipulation could promote the release of tumour cells into the bloodstream, but the ligation of pulmonary vein before the ligation of the pulmonary artery may partly prevent such release during surgery.

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.

Disulphide reaction staining for the identification of integumental elastic fibres

Based on specific methods (Sippel-APM-chromotropic acid technique; IC3-PE-maleimide fluorescence reaction) and skin samples of four domesticated mammals (dog, cattle, horse, pig), disulphide groups were demonstrated in the elastic component of the basement membrane of the epidermis, the elastic fibre system of the dermis, the elastic components of the connective tissue sheath of hair follicles, apocrine tubular glands, and sebaceous glands, and of the connective tissue surrounding the cutaneous muscle. The results are discussed regarding the relation of this reaction staining to the presence of microfibrils (fibrillin) in the elastic fibres.

Microbial detection in microfluidic devices through dual staining of quantum dots-labeled immunoassay and RNA hybridization

This paper reported the development of a microfludic device for the rapid detection of viable and nonviable microbial cells through dual labeling by fluorescent in situ hybridization (FISH) and quantum dots (QDs)-labeled immunofluorescent assay (IFA). The coin sized device consists of a microchannel and filtering pillars (gap = 1–2 μm) and was demonstrated to effectively trap and concentrate microbial cells (i.e. Giardia lamblia). After sample injection, FISH probe solution and QDs-labeled antibody solution were sequentially pumped into the device to accelerate the fluorescent labeling reactions at optimized flow rates (i.e. 1 and 20 μL/min, respectively). After 2 min washing for each assay, the whole process could be finished within 30 min, with minimum consumption of labeling reagents and superior fluorescent signal intensity. The choice of QDs 525 for IFA resulted in bright and stable fluorescent signal, with minimum interference with the Cy3 signal from FISH detection.

Multiple Alternative Splicing and Differential Expression of dmrt1 During Gonad Transformation of the Rice Field Eel1

Morphologically distinct males and females are observed throughout the animal kingdom. Why and how sex evolved and is maintained in most living organisms remains a key question in cellular and evolutionary biology. Here we report that four isoforms of dmrt1 (dsx- and mab3-related transcription factor 1) are generated in testis, ovotestis, and ovary by alternative splicing in the rice field eel, a fresh water fish that undergoes natural sex reversal from female to male during its life cycle. These transcripts encode four different size proteins with 301, 196, 300, and 205 amino acids. Like fly doublesex splicing, the dmrt1 of the rice field eel is also alternatively spliced at the 3' region, which generates diverse isoforms in gonads by alternative use of 3' sequences. Not only is dmrt1 expressed specifically in gonads, but its multiple isoforms are differentially coexpressed in gonadal epithelium during gonad transformation. Expression levels of a and b isoforms of dmrt1 ranged from low to high (ovary < ovotestis I < ovotestis II < ovotestis III < testis), based on comparisons of mean values from real-time fluorescent quantitative reverse transcription-polymerase chain reaction analysis. The overall expression level of dmrt1 b was much lower than that of dmrt1 a. Expression of dmrt1 d was not only low, but it also did not change significantly during sex transformation. The differential expression of dmrt1 isoforms may also be regulated by their 3' untranslated regions (UTRs), although these 3' UTRs do not contribute to intracellular localization of the Dmrt1 protein. These results provide new insight into roles of regulation at the level of splicing of dmrt1 in governing the sex differentiation cascade.

Analysis of Gene Regulation in Rabbit Corneal Epithelial Cells Induced by Ultraviolet Radiation

Ultraviolet (UV)-induced cataracts are becoming a major environmental health concern because of the possible decrease in the stratospheric ozone layer. Experiments were designed to isolate gene(s) affected by UV irradiation in rabbit cornea tissues using fluorescent differential display-reverse transcription-polymerase chain reaction (FDDRT-PCR). The epithelial cells were grown in standard medium for 2 or 4 hours post treatment. Cornea epithelial cells were irradiated with UVB for 20 minutes. RNA was extracted and amplified by reverse transcriptase-polymerase chain reaction using poly A+ specific anchoring primers and random arbitrary primers. Polyacrylamide gel electrophoresis revealed several differentially expressed genes in untreated versus UV irradiated cells. Complimentary DNA (cDNA) fragments resulting from fluorescent differentially expressed mRNAs were eluted from the gel and re-amplified. The re-amplified PCR products were cloned directly into the PCR-TRAP cloning system. These data showed that FDDRT-PCR is a useful technique to elucidate UV-regulated gene expressions. Future experiments will involve sequence analysis of cloned inserts. The identification of these genes through sequence analysis could lead to a better understanding of cataract formation via DNA damage and mechanisms of prevention.

2002 W.A.E. McBryde Award Lecture — Affinity recognition, capillary electrophoresis, and laser-induced fluorescence polarization for ultrasensitive bioanalysis

The combination of affinity recognition, capillary electrophoresis (CE), laser-induced fluorescence (LIF), and fluorescence polarization for the ultrasensitive determination of compounds of biological interest is described. Competitive immunoassays using CE–LIF eliminate the need for fluorescently labeling trace analytes of interest and are particularly useful for determination of small molecules, such as cyclosporine, gentamicin, vancomycin, and digoxin. Fluorescence polarization allows for differentiation of the antibody-bound from the unbound small molecules. Noncompetitive affinity CE–LIF assays are shown to be highly effective in the determination of biomarkers for DNA damage and HIV-1 infection. An antibody (or aptamer) is used as a fluorescent probe to bind with a target DNA adduct (or the reverse transcriptase of the HIV-1 virus), with the fluorescent reaction products being separated by CE and detected by LIF. Aptamers are attractive affinity probes for protein analysis because of high affinity, high specificity, and the potential for a wide range of target proteins. Fluorescence polarization provides unique information for studying molecular interactions. Innovative integrations of these technologies will have broad applications ranging from cancer research, to biomedical diagnosis, to pharmaceutical and environmental analyses.Key words: capillary electrophoresis, laser-induced fluorescence, fluorescence polarization, immunoassay, affinity probes, antibodies, aptamers, DNA damage, toxins, therapeutic drugs.

Spectrofluorimetric Determination of Warfarin Sodium by Using N1-Methylnicotinamide Chloride as a Fluorigenic Agent

A spectrofluorimetric method is described for the determination of drugs containing active methylene groups adjacent to carbonyl groups. The method was applied successfully to the determination of warfarin sodium in laboratory-prepared mixtures, in commercial tablets, and in spiked human plasma samples. Finally, the method was applied to the determination of the steady-state concentration of warfarin sodium in the blood of a hospitalized patient. The method involves the reaction of warfarin sodium with 0.2 ml (0.4 x 10(-3)M) N1-methylnicotinamide chloride reagent in the presence of 3 mL 1.0N NaOH and cooling in ice for 8 min, followed by adjustment of the pH to 2.0, using formic acid and heating for 4 min, whereby a highly fluorescent reaction product is produced. The optimal wavelengths of excitation and emission were determined by using a synchronous wavelength search and found to be 284 and 354 nm, respectively. The standard curves were linear over a concentration range of 50-1500 ng/mL in both aqueous solutions and spiked human plasma samples. The mean recoveries (+/- standard deviation) were 101.157 (+/-1.33) and 95.73 (+/-1.88%) for aqueous solutions and spiked human plasma samples, respectively. The method showed good specificity and precision. The proposed method is simple and economical because of its minimal instrumentation and chemicals requirements. Nevertheless, it is highly sensitive, specific, and reproducible. Accordingly, it is suitable for quality-control applications, drug monitoring, and bioavailability and bioequivalency studies.