Chemiluminescence determination of citrate and pyruvate and their mixtures by the stopped-flow mixing technique

A novel kinetic chemiluminescent method using the stopped-flow mixing technique has been investigated for the rapid and sensitive determination of citrate and pyruvate. The method is based on a tris(2,2′-bipyridiyl)ruthenium(III) (Ru(bpy)₃³⁺) chemiluminescence (CL) reaction. Ru(bpy)₃³⁺ was generated in the mixing chamber by oxidising tris(2,2′-bipyridyl)ruthenium(II) with cerium(IV). After selecting the best operating parameters, calibration graphs were obtained over the concentration ranges 0.38-38 μg ml⁻¹ and 8.7-1300 ng ml⁻¹ for citrate and pyruvate, respectively. The limits of detection were 0.1 μg ml⁻¹ for citrate and 0.3 ng ml⁻¹ for pyruvate. Based on the differential rate of the chemiluminescent reaction corresponding to citrate and pyruvate, a very simple kinetic procedure was developed for the simultaneous determination of both compounds. Mixtures of citrate and pyruvate in ratios between 15:1 and 1.5:1 were satisfactorily resolved. The proposed method was successfully applied to the determination of citrate in pharmaceutical formulations, pyruvate in animal blood serum and both compounds in human urine.     

Flow injection chemiluminescence determination of paracetamol

A simple chemiluminometric method using flow injection has been developed for the determination of paracetamol (acetaminophen), based on the chemiluminescence produced by the reduction of tris(2,2′-bipyridyl)ruthenium(III). The latter is obtained by oxidation of tris(2,2′-bipyridyl)ruthenium(II) by potassium permanganate in dilute sulphuric acid in the presence of paracetamol. A standard or sample solution was injected into the ruthenium(II) stream (flow rate 1.5 ml min⁻¹) which was then merged with potassium permanganate in dilute sulphuric acid stream (flow rate 0.5 ml min⁻¹). The chemiluminescence intensity is enhanced by the presence of manganese(II) ions. Under the optimum conditions, a linear calibration graph was obtained over the range of 0.3-50.0 μg ml⁻¹ and the detection limit was 0.2 μg ml⁻¹ (s/n = 3). The relative standard deviation of the proposed method calculated from 20 replicate injections of 5.0 μg ml⁻¹ paracetamol was 1.1%. The sample throughput was 90 h⁻¹. The method was successfully applied to the determination of paracetamol in commercial pharmaceutical formulations.     

Flow-injection chemiluminescence assay for ultra-trace determination of DNA using rhodamine B-Ce(IV)-DNA ternary system in sulfuric acid media

A novel flow-injection chemiluminescence method for the determination of DNA at ultra-trace level has been established. In 0.8 M sulfuric acid media, the chemiluminescence of the rhodamine B-cerium (IV) or Ce(IV) system is enhanced by DNA, activated previously by imidazole-HCl buffer solution (pH 7.0). The enhanced intensity of chemiluminescence is in proportion to log DNA concentration 1.0×10⁻⁸ to 0.1 μg ml⁻¹ for herring sperm DNA and 2.0×10⁻⁶ to 0.2 μg ml⁻¹ for calf thymus DNA with 3σ detection limits of 8.3×10⁻⁹ μg ml⁻¹ for herring sperm DNA and 3.5×10⁻⁷ μg ml⁻¹ for calf thymus DNA, respectively. The relative standard deviation for 1.0×10⁻⁴ μg ml⁻¹ herring sperm DNA was 0.99% and 2.0×10⁻³ μg ml⁻¹ for calf thymus DNA was 1.1% (n=11). Using the optimized system, DNA contents in six synthetic samples has been determined with recoveries of 99.5-109.0%. The possible mechanism has also been studied in this paper.     

Ultra-trace level determination of hydroquinone in waste photographic solutions by UV-vis spectrophotometry

A simpler UV-vis spectrophotometric method was investigated for hydroquinone (HQ) determination using KMnO₄ as oxidizing agent for conversion of HQ to p-benzoquinone (BQ) as well as signal enhancer. Various parameters such as analytical wavelength, stability time, temperature, pH, solvent effect and interference of chemicals were checked and parameters optimized by using 1 μg ml⁻¹ standard solution of HQ. Beer's Law was applicable in the range of 0.07-2 μg ml⁻¹ and 0.005-0.05 μg ml⁻¹ at 245.5 nm and at 262 nm for aqueous standard solutions of HQ with linear regression coefficient value of 0.9978 and 0.9843 and detection limit of 0.021 μg ml⁻¹ and 0.0016 μg ml⁻¹ HQ, respectively. Standard deviation of 1.7% and 2.4% was true for 1 μg ml⁻¹ and 0.03 μg ml⁻¹ HQ solution (n = 11) run at respective wavelengths. The method was successfully applied to dilute waste photographic developer samples for free HQ determination.     

Determination of thyroxine hormone by luminol chemiluminescence

A chemiluminescence (CL) flow system for determination of thyroxine (Thy) is presented. It is based on the catalytic effect of cobalt(II) on the CL reaction between luminol and hydrogen peroxide. The iodinated chemical structure of Thy causes a heavy atom effect. The luminol CL signals show significant quenching by Thy. The calibration graph for Thy is linear for 15-70 μg ml⁻¹ and the 3σ detection limits are 27 μg ml⁻¹ for d-Thy and 23 μg ml⁻¹ for l-Thy.     

Spectrofluorimetric determination of iridium(IV) traces using 4-pyridone derivatives

A new spectrofluorimetric determination of iridium(IV) with 3-hydroxy-2-methyl-1-phenyl-4-pyridone (HX) or 3-hydroxy-2-methyl-1-(4-tolyl)-4-pyridone (HY) is reported. Iridium(IV) react with HX or HY and chelates were extracted into chloroform or dichloromethane. The organic phase showed fluorescence. The fluorescence measurements to quantify iridium were carried out in its fluorescent band centred at λ_ex=373 nm and λ_em=480 nm. Under optimal conditions, the calibration graphs were linear over the concentration range of 0.1-7.6 μg ml⁻¹ of iridium for Ir(IV)-HX and 0.1-5.8 μg ml⁻¹ for Ir(IV)-HY with a correlation coefficients of 0.999 and 0.992 and relative standard deviation of ±1.1%.The method is free from interference by Rh(III) and Pt(IV), which normally interfere with other methods. Iridium can be determined in the presence of 300-fold excess of rhodium(III) and 10-fold excess of platinum(IV).The method was applied successfully to the determination of iridium in some synthetic mixtures and mineral sample gave satisfactory results.     

Chemiluminescence determination of ultramicro DNA with a flow-injection method

A high sensitive flow-injection chemiluminescence method for determination of calf thymus DNA and herring sperm DNA has been developed. The method is based on the chemiluminescence reaction of Rhodamine B-cerium(IV)-thermally denatured DNAs in sulfuric acid media. The proposed procedure allows quantitation of DNAs in the range 2.6×10⁻⁵ to 0.26 μg ml⁻¹ for calf thymus DNA and 5.0×10⁻⁸ to 5.0×10⁻⁵ μg ml⁻¹ for herring sperm DNA with correlation coefficients 0.9998 and 0.9996 (both n=11), respectively. The detection limits (3σ) are 6.5×10⁻⁶ μg ml⁻¹ for calf thymus DNA and 4.3×10⁻⁸ μg ml⁻¹ for herring sperm DNA. The possible mechanism of chemiluminescence in the system is discussed.     

Flow-injection chemiluminescent determination of metoclopramide hydrochloride in pharmaceutical formulations and biological fluids using the [Ru(dipy)(3)(2+)]-permanganate system

A flow-injection (FI) methodology using (2,2′-dipyridyl) ruthenium(II) [Ru(dipy)₃²⁺] chemiluminescence (CL) was developed for the rapid and sensitive determination of metoclopramide hydrochloride. The method is based on the CL reaction of metoclopramide with Ru(dipy)₃²⁺ and KMnO₄ in a sulfuric acid medium. Under the optimum conditions, a calibration graph was obtained over the concentration range 0.005-3.5 μg ml⁻¹ with a limit of detection (S/N=2) of 1 ng ml⁻¹. The correlation coefficient was 0.99993 (n=8) with a relative standard deviation of 0.48% for 10 determinations of 1 μg ml⁻¹ of drug. The method was successfully applied to the determination of metoclopramide in pharmaceutical preparations and biological fluids after IP administration of 25 mg kg⁻¹ dose to rats. The elimination half-life was 2.5±0.4 h.     

Flow-injection chemiluminescence determination of dihydralazine sulfate based on hexacyanoferrate(III) oxidation sensitized by eosin Y

A novel flow-injection chemiluminescence (CL) method for the determination of dihydralazine sulfate (DHZS) is described. The method is based on the reaction between DHZS and hexacyanoferrate(III) in alkaline solution to give weak CL signal, which is dramatically enhanced by eosin Y. The CL emission allows quantitation of DHZS concentration in the range 0.02-2.8 μg ml⁻¹ with a detection limit (3σ) of 0.012 μg ml⁻¹. The experimental conditions for the CL reaction are optimized and the possible reaction mechanism is discussed. The method has been applied to the determination of DHZS in pharmaceutical preparations and compared well with the high performance liquid chromatography (HPLC) method.     

Simultaneous determination of 11 drugs belonging to four different groups in human urine samples by reversed-phase high-performance liquid chromatography method

A new, accurate, sensitive and fast reversed-phase high-performance liquid chromatography (RP-HPLC) as an analytical method for the quantitative determination of 11 drugs in human urine was worked out, optimized and validated. The objects of analysis were imipenem (IMP), paracetamol (PAR), dipyrone (DPR), vancomycin (VCM), amikacin (AMK), fluconazole (FZ), cefazolin (CFZ), prednisolone (PRE), dexamethasone (DEX), furosemide (FUR) and ketoprofen (KET) belonging to four different groups (antibiotics, analgesic, demulcent and diuretic). For HPLC analysis, diode array (DAD) and fluorescence (FL) detectors were used. The separation of analyzed compounds was conducted by means of a LiChroCART^® Purospher^® C₁₈e (125 mm × 3 mm, particle size 5 μm) analytical column with LiChroCART^® LiChrospher^® C₁₈ (4 mm × 4 mm, particle size 5 μm) pre-column with gradient elution. Analyzed drugs were determined within 20 min. The mobile phase was comprised of various proportions of methanol, acetonitrile and 0.05% trifluoroacetic acid in water. AMK was separated and determined from human urine using ortho-phthaldialdehyde-3-mercaptopropionic acid (OPA-3-MPA) as a fluorescent reagent by RP-HPLC-FL. The following retention times for drugs IMP, PAR, DPR, VCM, AMK, FZ, CFZ, PRE, DEX, FUR and KET in human urine were found: 4.01 min, 4.86 min, 6.71 min, 8.14 min, 9.46 min, 10.01 min, 10.90 min, 13.34 min, 14.06 min, 16.03 min and 18.98 min, respectively. Excellent linearity was obtained for compounds in the range of concentration: 0.35-42 μg ml⁻¹, 0.5-45 μg ml⁻¹, 4.5-38 μg ml⁻¹, 0.25-25 μg ml⁻¹, 0.5-35 μg ml⁻¹, 0.25-22 μg ml⁻¹, 0.03-52 μg ml⁻¹, 0.15-25 μg ml⁻¹, 0.25-28 μg ml⁻¹, 0.05-18 μg ml⁻¹ and 0.15-35 μg ml⁻¹ for IMP, PAR, DPR, VCM, AMK, FZ, CFZ, PRE, DEX, FUR and KET, respectively. The limits of detection (LOD) and limits of quantification (LOQ) for analyzed drugs were calculated in all cases and recovery studies were also performed. Ten human urine samples obtained from patients treated in hospital have been tested. In analyzed samples, one or more drugs from the 11 examined drugs were detected. The concentrations of examined drugs in urine samples ranged between: 1.5-12 μg ml⁻¹ of PAR, 5.2-11.5 μg ml⁻¹ of DPR, 0.13-9.5 μg ml⁻¹ of CFZ and 0.1-8 μg ml⁻¹ of FUR. This method can be successfully applied to routine determination of all these drugs in human urine samples.     

Sensitive and selective determination of molybdenum by flow injection chemiluminescence method combined with controlled potential electrolysis technique

A sensitive and selective flow injection chemiluminescence (CL) method combined with controlled potential electrolysis technique was described for the determination of molybdenum. The method is based on the chemiluminescence reaction of luminol with unstable molybdenum(III) in alkaline solution. The molybdenum(III) was on-line reduced from molybdenum(VI) via controlled potential electrolysis technique using a homemade flow-through carbon electrolytic cell at the potential of −0.6 V (versus Ag/AgCl). The method allows the determination of molybdenum in the 5.0×10⁻¹⁰ to 5.0×10⁻⁷ g ml⁻¹ range with a limit of detection (3σ) of 5×10⁻¹¹ g ml⁻¹ molybdenum. The relative standard deviation is 2.6% for the 1.0×10⁻⁹ g ml⁻¹ molybdenum solution in 11 repeated measurements. This method was successfully applied to the determination of molybdenum in water samples.     

Synchronous fluorescence determination of protein with functionalized CdS nanoparticles as a fluorescence probe

Nanometer-sized fluorescent particles have been successfully synthesized. A synchronous fluorescence method, with high sensitivity and selectivity, has been developed for rapid determination of protein with functionalized CdS as a fluorescence probe. When Δλ=260 nm, maximum synchronous fluorescence is produced at 274 nm at pH 7.0. Under optimal conditions, the calibration graphs are linear over the range 0.1-3.0 μg ml⁻¹ for bovine serum albumin (BSA), 0.1-11.0 μg ml⁻¹ for γ-globulin (γ-G) and 0.1-1.4 μg ml⁻¹ for human serum albumin (HSA), respectively. Limits of determination were 0.01 μg ml⁻¹ for BSA, 0.019 μg ml⁻¹ for γ-G and 0.021 μg ml⁻¹ for HSA, respectively. The relative standard deviations of seven replicate measurements were 1.8% for 1.0 μg ml⁻¹ BSA, 2.2% for 1.0 μg ml⁻¹ γ-G and 2.3% for 1.0 μg ml⁻¹ HSA.     

Determination of ternary mixtures of penicillin G, benzathine and procaine by liquid chromatography and factorial design study

In the present work, a rapid and sensitive method for simultaneous determination of penicillin G (PG), benzathine (BE) and procaine (PR) in drug and serum media is introduced. The polar hydro-organic (55/45) mobile phases containing an aqueous solution adjusted to pH = 3.7 and an organic solvent (MeOH) including triethylamine (TEA) and trifluroacetic acid (TFA) are used. The flow rate of 1 ml min⁻¹, a C₈ column (150 mm × 46 mm) with 5 μm i.d. and wavelength at 215 nm are selected for optimal separation condition. The limit of detection (LOD), linear concentration range and relative standard deviation (R.S.D.) of this method for the PG are 1.1 μg ml⁻¹, 10-2400 μg ml⁻¹ and 1.7% and for the BE are 1.2 μg ml⁻¹, 12-2100 μg ml⁻¹ and 1.8% and for the PR are 1.5 μg ml⁻¹, 20-2000 μg ml⁻¹ and 2%, respectively. The factorial design is used for the determination of main and interaction effects of pH, flow rate and concentration of MeOH, TEA and TFA in the separation at two levels. Also, the analysis of variance (ANOVA) table is obtained. The results show that TFA and TEA have higher effect than concentration of MeOH, pH and flow rate factors.     

Chemiluminescence determination of carbofuran at trace levels in lettuce and waters by flow-injection analysis

A simple, fast chemiluminescence (CL) flow-injection (FI) method based on the reaction of luminol with KMnO₄ in alkaline medium has been described for the direct determination of carbofuran. The method is based on the enhancing effect in the emission light from the oxidation of luminol produced in presence of carbofuran. The optimisation of instrumental and chemical variables influencing the CL response of the method has been carried out by applying experimental design, using the proposed flow-injection manifold. Under the optimal conditions, the CL intensity was linear for a carbofuran concentration over the range of 0.06-0.5 μg ml⁻¹, with a detection limit of 0.02 μg ml⁻¹. The method has been successfully applied to the determination of carbofuran residues in spiked water and lettuce samples.     

Sequential flow injection analysis of ammonium and nitrate using gas phase molecular absorption spectrometry

A flow injection method on the basis of gas phase molecular absorption is described for the sequential determination of ammonium and nitrate. Two hundred microliters of sample solution is injected into the flow line. For ammonium determination, the sample zone is directed to a line in which reacts with NaOH (13 M) and produces ammonia. But for nitrate determination, the sample zone is passed through the on-line copperized zinc (Zn/Cu) reduction column and produces ammonium ion and in the follows ammonia. The produced ammonia in both cases is purged into the stream of N₂ carrier gas. The gaseous phase is separated from the liquid phase using a gas-liquid separator and then is swept into a flow through cell, which has been positioned in the cell compartment of an UV-Vis spectrophotometer. The absorbance of the gaseous phase is measured at 194 nm. Under selected conditions for sequential analysis of ammonium and nitrate, linear relations were found between the peak heights of absorption signals and concentrations of ammonium (10-650 μg ml⁻¹) and nitrate (20-800 μg ml⁻¹). The limit of detections for ammonium and nitrate analysis were 8 and 10 μg ml⁻¹, respectively. The relative standard deviations of repeated measurements of 50 μg ml⁻¹ of ammonium and nitrate were 2.0, 2.9%, respectively. Maximum sampling rate was about 40 samples/h. The method was applied to the determination of ammonium in pharmaceutical products and the sequential determination of ammonium and nitrate in spiked water samples.     

Micro determination of ascorbic acid using methyl viologen

A new simple and sensitive analytical spectrophotometric method is developed for the determination of ascorbic acid reduces methyl viologen to form a stable blue coloured free radical ion. This method has a sensitivity and lower limit detection of 0.1 μg ml⁻¹ of ascorbic acid (0.1 ppm) which is comparable to the flow injection analysis reported earlier. Beer's law is obeyed over the concentration range of 1.0-10 μg ml⁻¹ of ascorbic acid per 10 ml of the final solution (0.1-1.0 μg ml⁻¹) at 600 nm. The molar absorptivity and Sandell's sensitivity were found to be 1.5 × 10⁵ ± 100 l mol⁻¹ cm⁻¹ and 0.001 μg cm⁻², respectively. The method has been applied to the determination of ascorbic acid in food, pharmaceuticals and biological samples.     

Fluorescence and terbium-sensitised luminescence determination of garenoxacin in human urine and serum

Fluorescence and terbium-sensitised luminescence properties of new quinolone garenoxacin have been studied. The fluorimetric method allows the determination of 0.060-0.600 μg ml⁻¹ of garenoxacin in aqueous solution containing HCl/KCl buffer (pH 1.5) with λ_exc=282 nm and λ_em=421 nm. Micellar-enhanced fluorescence was also studied, leading to a higher than 400% increase in analytical signal in presence of 12 mM sodium dodecyl sulphate (SDS), allowing the determination of 0.020-0.750 μg ml⁻¹ of garenoxacin. The terbium-sensitised luminescence method allows the determination of 0.100-1.500 μg ml⁻¹ of garenoxacin in 12 mM SDS solution containing 0.08 M acetic acid/sodium acetate buffer (pH 4.1) and 7.5 mM Na₂SO₃ (chemical deoxygenation agent), with λ_exc=281 nm and λ_em=546 nm. Relative standard deviation (R.S.D.) values for the three methods were in the range 1.0-2.0%. The proposed procedures have been applied to the determination of garenoxacin in spiked human urine and serum.     

Flow injection chemiluminescence determination of carbaryl using photolytic decomposition and photogenerated tris (2,2′-bipyridyl)ruthenium(III)

A flow injection (FI) system combined with two photochemical processes is developed for the sensitive and rapid determination of carbaryl. It is based on the on-line photo-conversion of carbaryl into methylamine which subsequently reacts with Ru(bpy)₃³⁺ generated through the on-line photo-oxidation of Ru(bpy)₃²⁺ with peroxydisulphate. The linear concentration range of application was 0.04-4.0 μg ml⁻¹ of carbaryl, with an R.S.D. of 1.2% (for a level of 0.50 μg ml⁻¹) and a detection limit of 0.012 μg ml⁻¹. The sample throughput was 200 injections per hour. The applicability of the method was demonstrated by determining carbaryl in commercial formulations, water, soil, grain and blood serum.     

Non-equilibrium determination of metoclopramide and tetracaine hydrochloride by sequential injection spectrophotometry

A new sequential injection spectrophotometric method was proposed for the determination of metoclopramide and tetracaine hydrochloride. The method was based on the detection of an unstable red intermediate compound resulting from the reaction of metoclopramide or tetracaine hydrochloride with potassium dichromate, in the presence of sodium oxalate, in sulfuric acid solution. The related reaction mechanisms of this new method have been studied. The experimental conditions were optimized for the stopped-flow and continuous-flow sequential injection models. For continuous flow, the linear range for determination of metoclopramide, the detection limit and the sampling frequency were 13-130 μg ml⁻¹, 9.4 μg ml⁻¹ and 40 samples per hour, respectively. For stopped flow, they were 3-42 μg ml⁻¹, 1.0 μg ml⁻¹ and 18 h⁻¹, respectively. Adopting the continuous-flow model for tetracaine hydrochloride, the linear range was 25-300 μg ml⁻¹, and the detection limit was 18.0 μg ml⁻¹ with sampling frequency of 40 h⁻¹. This method has been used to determine metoclopramide and tetracaine hydrochloride in pharmaceutical preparations, and the results are compared with those determined by the pharmacopoeia method. Statistical analysis reveals that there was no evidence of significant difference between the methods.     

Development of an optical chemical sensor based on 2-(5-bromo-2-pyridylazo)-5-(diethylamino)phenol in Nafion for determination of nickel ion

An optical chemical sensor based on immobilization of 2-(5-bromo-2-pyridylazo)-5-(diethylamino)phenol (Br-PADAP) in Nafion membrane is described. The membranes were cast onto glass substrates and were used for the determination of nickel in aqueous solutions by spectrophotometry. The sensor system is highly transparent, mechanically stable and showed no evidence of reagent leaching. The influence of several parameters such as pH, ligand concentration, and type and concentration of regenerating solution were optimized. The sensor system showed good sensitivity in the range 0.5-20 μg ml⁻¹ with a detection limit of 0.3 μg ml⁻¹ Ni(II). The sensor has been incorporated into a home-made flow-through cell for determination of nickel in flowing streams with improved sensitivity, precision and detection limit. The calibration curve in the flow system was linear in the range 0.1-16 μg ml⁻¹ with a detection limit of 0.07 μg ml⁻¹. The sensor is easily regenerated by dilute nitric acid solution. The proposed method was successfully applied to the determination of nickel content in vegetable oil and chocolate samples and the results were compared with those obtained using atomic absorption spectrometry.