Normal spectrophotometric and stopped-flow spectrofluorimetric sequential injection methods for the determination of alendronic acid, an anti-osteoporosis amino-bisphosphonate drug, in pharmaceuticals

A normal spectrophotometric and a stopped-flow (SF) spectrofluorimetric method have been developed and optimized for the determination of alendronic acid (ALD) in its pharmaceutical formulations. Both methods are automated using the sequential injection analysis (SIA) principle. The spectrophotometric assay is based on the reaction of the analyte with Cu(II) ions in acidic medium to form an UV-absorbing derivative (λ_max = 240 nm). The SF spectrofluorimetric method is based on the reaction of ALD with o-phthalaldehyde (OPA) in the presence of 2-mercaptoethanol at basic medium (λ_ex = 340 nm/λ_em = 455 nm). Linear calibration curves were obtained in the range 1.0-60.0 mg l⁻¹ ALD for the UV method, and in the range 0.13-10.0 mg l⁻¹ ALD for the SF spectrofluorimetric one. The sampling rates were 60 and 30 h⁻¹, respectively. The developed assays are critically compared and their advantages are discussed. Both methods were applied to the analysis of an ALD containing pharmaceutical formulation with satisfactory accuracy and precision.     

The inner filter effects and their correction in fluorescence spectra of salt marsh humic matter

The inner filter effects in synchronous fluorescence spectra (Δλ = 60 nm) of sedimentary humic substances from a salt marsh were studied. Accordingly to their type and the influence of plant colonization, these humic substances have different spectral features and the inner filter effects act in a different manner. The fluorescence spectra of the humic substances from sediments with colonizing plants have a protein like band (λ_exc = 280 nm) which is strongly affected by primary and secondary inner filter effects. These effects were also observed for the bands situated at longer wavelengths, i.e., at λ_exc = 350 nm and λ_ex = 454 nm for the fulvic acids (FA) and humic acids (HA), respectively. However, they are more important for the band at 280 nm, causing spectral distortions which can be clearly seen when the spectra of solutions 40 mg L⁻¹ of different samples (Dissolved Organic Carbon – DOC ∼ 20 mg L⁻¹) are compared with and without correction of the inner filter effects. The importance of the spectral distortions caused by inner filter effects has been demonstrated in solutions containing a mixture of model compounds which represent the fluorophores detected in the spectra of sedimentary humic samples. The effectiveness of the mathematical correction of the inner filter effects in the spectra of those solutions and of solutions of sedimentary humic substances was studied. It was observed that inner filter effects in the sedimentary humic substances spectra can be mathematically corrected, allowing to obtain a linear relationship between the fluorescence intensity and humic substances concentration and preventing distortions at concentrations as high as 50 mg L⁻¹ which otherwise would obscure the protein like band.     

Hybrid sequential injection-flow injection manifold for the spectrophotometric determination of total sulfite in wines using o-phthalaldehyde and gas-diffusion

A new automated spectrophotometric method for the determination of total sulfite in white and red wines is reported. The assay is based on the reaction of o-phthalaldehyde (OPA) and ammonium chloride with the analyte in basic medium under SI conditions. Upon on-line alkalization with NaOH, a blue product is formed having an absorption maximum at 630 nm. The parameters affecting the reaction - temperature, pH, ionic strength, amount concentration and volume of OPA, amount concentration of ammonium chloride, flow rate and reaction coil length - and the gas-diffusion process - sample and HCl volumes, length of mixing coil, donor flow rate - were studied. The proposed method was validated in terms of linearity (1-40 mg L⁻¹, r = 0.9997), limit of detection (c_L = 0.3 mg L⁻¹) and quantitation (c_Q = 1.0 mg L⁻¹), precision (s_r = 2.2% at 20 mg L⁻¹ sulfite, n = 12) and selectivity. The applicability of the analytical procedure was evaluated by analyzing white and red wine samples, while the accuracy as expressed by recovery experiments ranged between 96% and 106%.     

Reversed flow injection spectrophotometric determination of chlorate

An interfacing has been developed to connect a spectrophotometer with a personal computer and used as a readout system for development of a simple, rapid and sensitive reversed flow injection (rFI) procedure for chlorate determination. The method is based on the oxidation of indigo carmine by chlorate ions in an acidic solution (dil. HCl) leading to the decrease in absorbance at 610 nm. The decrease in absorbance is directly related to the chlorate concentration present in the sample solutions. Optimum conditions for chlorate were examined. A linear calibration graph over the range of 0.1-0.5 mg L⁻¹ chlorate was established with the regression equation of Y = 104.5X + 1.0, r² = 0.9961 (n = 6). The detection limit (3σ) of 0.03 mg L⁻¹, the limit of quantitation (10σ) of 0.10 mg L⁻¹ and the RSD of 3.2% for 0.3 mg L⁻¹ chlorate (n = 11) together with a sample throughput of 92 h⁻¹ were obtained. The recovery of the added chlorate in spiked water samples was 98.5 ± 3.1%. Major interferences for chlorate determination were found to be BrO₃⁻, ClO₂⁻, ClO⁻ and IO₃⁻ which were overcome by using SO₃²⁻ (as Na₂SO₃) as masking agent. The method has been successfully applied for the determination of chlorate in spiked water samples with the minimum reagent consumption of 14.0 mL h⁻¹. Good agreement between the proposed rFIA and the reference methods was found verified by Student's t-test at 95% confidence level.     

A green analytical procedure for flow-injection determination of nitrate in natural waters

Nitrate determination in waters is generally carried out with cadmium filings and carcinogenic reagents or by reaction with phenolic compounds in highly concentrated sulfuric acid medium. In this work, it was developed a green analytical procedure for nitrate determination in natural waters based on direct spectrophotometric measurements in ultraviolet, using a flow-injection system with an anion-exchange column for separation of nitrate from interfering species. The proposed method employs only one reagent (HClO₄) in a minimum amount (equivalent to 18 μL concentrated acid per determination), and allowed nitrate determination within 0.50-25.0 mg L⁻¹, without interference of up to 200.0 mg L⁻¹ humic acid; 1.0 mg L⁻¹ NO₂⁻; 200.0 mg L⁻¹ PO₄³⁻; 75.0 mg L⁻¹ Cl⁻; 50.0 mg L⁻¹ SO₄²⁻ and 15.0 mg L⁻¹ Fe³⁺. The detection limit (99.7% confidence level) and the coefficient of variation (n = 20) were estimated as 0.1 mg L⁻¹ and 0.7%, respectively. The results obtained for natural water samples were in agreement with those achieved by the reference method based on nitrate reduction with copperized cadmium at the 95% confidence level.     

Evaluation of alternate lines of Fe for sequential multi-element determination of Cu, Fe, Mn and Zn in soil extracts by high-resolution continuum source flame atomic absorption spectrometry

The usefulness of the secondary line at 252.744 nm and the approach of side pixel registration were evaluated for the development of a method for sequential multi-element determination of Cu, Fe, Mn and Zn in soil extracts by high-resolution continuum source flame atomic absorption spectrometry (HR-CS FAAS). The influence of side pixel registration on the sensitivity and linearity was investigated by measuring at wings (248.325, 248.323, 248.321, 248.329, and 248.332 nm) of the main line for Fe at 248.327 nm. For the secondary line at 252.744 nm or side pixel registration at 248.325 nm, main lines for Cu (324.754 nm), Mn (279.482 nm) and Zn (213.875 nm), sample flow-rate of 5.0 mL min⁻¹ and calibration by matrix matching, analytical curves in the 0.2-1.0 mg L⁻¹ Cu, 1.0-20.0 mg L⁻¹ Fe, 0.2-2.0 mg L⁻¹ Mn, 0.1-1.0 mg L⁻¹ Zn ranges were obtained with linear correlations better than 0.998. The proposed method was applied to seven soil samples and two soil reference materials (IAC 277; IAC 280). Results were in agreement at a 95% confidence level (paired t-test) with reference values. Recoveries of analytes added to soil extracts containing 0.15 and 0.30 mg L⁻¹ Cu, 7.0 and 14 mg L⁻¹ Fe, 0.60 and 1.20 mg L⁻¹ Mn, 0.07 and 0.15 mg L⁻¹ Zn, varied within the 94-99, 92-98, 93-101, and 93-103% intervals, respectively. The relative standard deviations (n = 12) were 2.7% (Cu), 1.4% (Fe - 252.744 nm), 5.7% (Fe - 248.325 nm), 3.2% (Mn) and 2.8% (Zn) for an extract containing 0.35 mg L⁻¹ Cu, 14 mg L⁻¹ Fe, 1.1 mg L⁻¹ Mn and 0.12 mg L⁻¹ Zn. Detection limits were 5.4 μg L⁻¹ Cu, 55 μg L⁻¹ Fe (252.744 nm), 147 μg L⁻¹ Fe (248.325 nm), 3.0 μg L⁻¹ Mn and 4.2 μg L⁻¹ Zn.     

Sequential injection fluorimetric determination of Sn in juices of canned fruits

The present work describes the development of a fast and robust sequential injection fluorimetric procedure for the determination of Sn in juices of canned fruits. The developed automatic methodology is based on the complexation of Sn with 8-hydroxyquinoline-5-sulfonic acid (HQSA) to form a fluorimetric product (λ_exc = 354 nm; λ_em = 510 nm). The influence of dimethylsulfoxide (DMSO) and cetylpyridinium bromide (CPB) on the sensitivity of the fluorimetric determination was evaluated.Linear calibration plots were obtained for Sn concentrations between 1 and 10 mg L⁻¹, with a detection limit of 0.38 mg L⁻¹. In each analytical cycle 0.006 mg of HQSA and 0.47 mg of CPB were consumed and 1.5 mL of effluent was generated.The developed methodology was applied to the determination of Sn in juices of canned fruits and the results complied with those furnished by an electrothermal atomic absorption spectrometry comparison procedure, with relative deviations lower than 5.2%.The automatic procedure exhibited good precision (R.S.D. < 1.4%) and the sampling rate was about 70 determinations per hour.     

Flow-injection determination of hydrogen peroxide based on fluorescence quenching of chromotropic acid catalyzed with Fe(II)

Flow-injection analysis system (FIA system), which was based on Fe(II)-catalyzed oxidation of chromotropic acid with hydrogen peroxide, was developed for the determination of hydrogen peroxide. The chromotropic acid has a fluorescence measured at λ_em = 440 nm (emission wavelength) with λ_ex = 235 nm (excitation wavelength), and the fluorescence intensity at λ_em = 440 nm quietly decreased in the presence of hydrogen peroxide and Fe(II), which was caused by Fe(II)-catalyzed oxidation of chromotropic acid with hydrogen peroxide. By measuring the difference of fluorescence intensity, hydrogen peroxide (1.0 × 10⁻⁸-1.0 × 10⁻³ mol L⁻¹) could be determined by the proposed FIA system, whose analytical throughput was 40 samples h⁻¹. The relative standard deviation (RSD) was 1.03% (n = 10) for 4.0 × 10⁻⁸ mol L⁻¹ hydrogen peroxide. The proposed FIA technique could be applied to the determination of hydrogen peroxide in rain water samples.     

Sequential injection methodology for carbon speciation in bathing waters

A sequential injection method (SIA) for carbon speciation in inland bathing waters was developed comprising, in a single manifold, the determination of dissolved inorganic carbon (DIC), free dissolved carbon dioxide (CO₂), total carbon (TC), dissolved organic carbon and alkalinity. The determination of DIC, CO₂ and TC was based on colour change of bromothymol blue (660 nm) after CO₂ diffusion through a hydrophobic membrane placed in a gas diffusion unit (GDU). For the DIC determination, an in-line acidification prior to the GDU was performed and, for the TC determination, an in-line UV photo-oxidation of the sample prior to GDU ensured the conversion of all carbon forms into CO₂. Dissolved organic carbon (DOC) was determined by subtracting the obtained DIC value from the TC obtained value. The determination of alkalinity was based on the spectrophotometric measurement of bromocresol green colour change (611 nm) after reaction with acetic acid. The developed SIA method enabled the determination of DIC (0.24–3.5 mg C L⁻¹), CO₂ (1.0–10 mg C L⁻¹), TC (0.50–4.0 mg C L⁻¹) and alkalinity (1.2–4.7 mg C L⁻¹ and 4.7–19 mg C L⁻¹) with limits of detection of: 9.5 μg C L⁻¹, 20 μg C L⁻¹, 0.21 mg C L⁻¹, 0.32 mg C L⁻¹, respectively. The SIA system was effectively applied to inland bathing waters and the results showed good agreement with reference procedures.     

Multi-pumping flow system for the determination of dissolved orthophosphate and dissolved organic phosphorus in wastewater samples

A multi-pumping flow system (MPFS) for the spectrophotometric determination of dissolved orthophosphate and dissolved organic phosphorus in wastewater samples is proposed. The determination of orthophosphate is based on the vanadomolybdate method. In-line ultraviolet photo-oxidation is employed to mineralise organic phosphorus to orthophosphate prior to detection. A solenoid valve allows the deviation of the flow towards the UV-lamp to carry out the determination of organic phosphorus.Calibration was found to be linear up to 20 mg P L⁻¹, with a detection limit (3s_b/S) of 0.08 mg P L⁻¹, an injection throughput of 75 injections h⁻¹ and a repeatability (R.S.D.) of 0.6% for the direct determination of orthophosphate. On the other hand, calibration graphs were linear up to 40 mg P L⁻¹, with a detection limit (3s_b/S) of 0.5 mg P L⁻¹, an injection throughput of 11 injections h⁻¹ and a repeatability (R.S.D.) inferior to 2.3% for the procedures involving UV photo-oxidation.     

Selective stopped-flow sequential injection method for the spectrophotometric determination of titanium in dental implant and natural Moroccan phosphate rock

The present work reports the first sequential injection (SI) method for the spectrophotometric determination of Ti(IV). The method is based upon the reaction of Ti(IV) with chromotropic acid (CA) in acidic medium to form a water-soluble complex (λ_max = 420 nm). The chemical and instrumental variables of the system that affected the reaction were studied. Selectivity was greatly enhanced using ascorbic acid. A linear calibration graph was obtained in the range 0.2-10.0 mg l⁻¹ Ti(IV) at a sampling frequency of 24 h⁻¹. The precision was satisfactory (s_r = 1.5% at 5.0 mg l⁻¹ Ti(IV), n = 12) and the 3σ limit of detection, c_L, was 0.7 mg l⁻¹ (n = 10). The developed method proved to be adequately selective and was applied successfully to the analysis of real samples (dental implant and natural Moroccan phosphate rock) giving accurate results based on recovery studies (98-105%).     

A novel application of immobilization on membranes for the separation and spectrofluorimetric quantification of amiloride and furosemide in pharmaceutical samples

A new, simple and highly sensitive method for spectrofluorimetric determination of amiloride (AMI) and furosemide (FUR) in pharmaceuticals is presented. The proposed method is based on the separation of AMI from FUR by solid-phase extraction using a nylon membrane, followed by spectrofluorimetric determination of both drugs, on the solid surface and the filtered aqueous solution, respectively. AMI shows low native fluorescence, but its separation-preconcentration by immobilization (solid-phase extraction) on nylon membrane surface provides a considerable enhancement in fluorescence intensity. The fluorescence determination is carried out at λ_ex = 237, λ_em = 415 nm for FUR; and λ_ex = 365, λ_em = 406 nm for AMI. The calibration graphs are linear in the range 3.20 × 10⁻⁴ to 0.8 μg mL⁻¹and 1.33 × 10⁻³ to 4.0 μg mL⁻¹, for AMI and FUR, respectively, with a detection limit of 9.62 × 10⁻⁵ and 4.01 × 10⁻⁴ μg mL⁻¹ (S/N = 3). The commonly found excipients in commercial pharmaceutical formulations do not interfere. The developed method is successfully applied to the determination of both drugs in pharmaceutical formulations.     

Solvent extraction flow-injection manifold for the simultaneous spectrophotometric determination of free cyanide and thiocyanate ions based upon on-line masking of cyanides by formaldehyde

This study reports a sensitive solvent extraction flow-injection (FI) method for the simultaneous spectrophotometric determination of free cyanide and thiocyanate in human saliva and pralidoxime solutions. Cyanide and thiocyanate form colored (λ_max=540 nm) ternary complexes with copper and 2,2′-dipyridyl-2-quinolylhydrazone (DPQH) that are extractable into chloroform. The determination of thiocyanates in the presence of cyanides is accomplished after on-line masking of the latter with formaldehyde through a binary inlet static mixer (BISM). Total thiocyanates and cyanides are determined in a second run, without the use of the masking agent. The proposed method allows the determination of the analytes in the range of 0-4 mg l⁻¹ thiocyanates and 0-3 mg l⁻¹ cyanides, with the 3σ detection limits being 0.007 and 0.004 mg l⁻¹, respectively. The precision of the method (s_r<1.0% at 1 mg l⁻¹ CN⁻ or SCN⁻, n=12 in both cases) and the sampling rates were quite satisfactory (60 injections per hour). The method was applied to the analysis of human saliva and pralidoxime solutions and gave recoveries in the range of 98.0-102.2% for both analytes whereas the mean relative error was e_r=1.7%.     

Highly sensitive and selective spectrofluorimetric determination of metoclopramide hydrochloride in pharmaceutical tablets and serum samples using Eu ion doped in sol-gel matrix

A simple, sensitive and selective spectrofluorimetric method for the determination of Metoclopramide hydrochloride (MCP) is developed. The MCP can remarkably enhances the luminescence intensity of the Eu³⁺ ion doped in sol-gel matrix at λ_ex = 380 nm in DMSO at pH 8.7. The intensity of the emission band of Eu³⁺ ion doped in sol-gel matrix increases due to energy transfer from MCP to Eu³⁺ in the excited state. The enhancement of the emission band of Eu³⁺ ion doped in sol-gel matrix at 617 nm was found to be directly proportional to the concentration of MCP with a dynamic range of 5 × 10⁻⁹ − 1.0 × 10⁻⁶ mol L⁻¹ and detection limit of 2.2 × 10⁻¹¹ mol L⁻¹.     

Selective determination of cyanides by gas diffusion-stopped flow-sequential injection analysis and an on-line standard addition approach

A highly selective sequential injection (SI) method for the automated determination of weak-acid-dissociable cyanides is reported. The analytical procedure is based on the on-line reaction of the analyte with ninhydrin in carbonate medium to form a coloured product (λ_max = 510 nm). Cyanides are removed from sample matrix by acidification through a gas-diffusion step incorporated in the SI manifold. The effect of instrumental and chemical variables was studied. By adopting an on-line standard addition protocol, the sensitivity of the proposed method was enhanced drastically, without affecting the determination range. The assay was validated in terms of linearity (up to 200 μg L⁻¹), limit of detection (c_L = 2.5 μg L⁻¹), limit of quantitation (c_Q = 7.5 μg L⁻¹), precision (s_r < 2.5% at 100 μg L⁻¹) and selectivity. High tolerance against critical species such as sulfides and thiocyanates was achieved. The applicability of the method was demonstrated by analyzing tap and mineral water samples at levels below the limits established by international E.U. and U.S. organizations. The percent recoveries were satisfactory in all cases, ranging between 94.2 and 103.6%.     

Highly selective flow injection spectrophotometric determination of gold based on its catalytic effect on the oxidation of variamine blue by potassium iodate in aqueous N,N-dimethylformamide medium

A highly selective and simple flow injection method is reported for the determination of Au(III) in jewel samples. The method is based on the catalytic effect of Au(III) on the oxidation of 4-amino-4′-methoxydiphenylamine hydrochloride (Variamine Blue B base, VB) by KIO₃. The colored reaction product was monitored spectrophotometrically at 546 nm. A volume fraction of 40% N,N-dimethylformamide (DMF) greatly enhances the selectivity of the method. The chemical (pH and concentrations of reagents) and instrumental variables (sample injection volume, reagents flow rates, reaction coil length) affecting the determination were studied and optimized. Under the selected values, the analyte could be determined in the range of 0.1-12.0 mg L⁻¹ (r = 0.9997), at a sampling rate of 120 h⁻¹. The proposed assay was precise (s_r = 0.8% at 5.0 mg L⁻¹ Au(III), n = 12) and adequately sensitive with a 3σ limit of detection of 0.03 mg L⁻¹. The method was successfully applied to the analysis of jewel samples. The obtained results were favorably compared to flame atomic absorption spectrometry (FAAS) used as a reference method.     

Determination of various alcohols based on a new immobilized enzyme fluorescence capillary analysis

A novel method for the determination of ethanol in tequila based on the immobilized enzyme fluorescence capillary analysis (IE-EFCA) has been proposed. Alcohol dehydrogenase (ADH) was immobilized in inner surface of a capillary and an immobilized enzyme capillary bioreactor (IE-ECBR) was formed. After nicotinamide adenine dinucleotide (NAD⁺) as an oxidizer is mixed with alcohol sample solution, it was sucked into the IE-ECBR. The fluorescence intensity of the mixed solution in the IE-ECBR was detected at λ_ex = 350 nm and λ_em = 459 nm. The experimental conditions are as follows: The reaction time is 20 min; temperature is 40 °C; the concentrations of phosphate buffer solution (pH 7.5) and NAD⁺ are 0.1 mol L⁻¹ and 5 mmol L⁻¹, respectively; immobilization concentration of ADH is 10 U L⁻¹. The determination range of ethanol is 2.0-15.0 g L⁻¹ (F = 10.44C + 6.6002, r > 0.9958); its detection limit is 1.11 g L⁻¹; and relative standard deviation is 1.9%. IE-EFCA method is applicable for the determination of the samples containing alcohol in medicine, industry and environment.     

A new tool for inorganic nitrogen speciation study: simultaneous determination of ammonium ion, nitrite and nitrate by ion chromatography with post-column ammonium derivatization by Nessler reagent and diode-array detection in rain water samples

The paper presents a new method for a simultaneous determination of inorganic nitrogen species in the oxidized (NO₂⁻, NO₃⁻) and reduced (NH₄⁺) form in rain water samples. The method is based on a system of nitrogen species separation employing ion exchange and diode-array detection. The ions are separated in a strong ion-exchanger, nitrites and nitrates are determined directly at 208 and 205 nm, respectively, while the ammonium ions are determined in the column hold-up time after a post-column derivatization by the Nessler reagent, at 425 nm. The use of a diode-array detector permits a simultaneous identification of the inorganic nitrogen species in 8 min. The detection limits obtained are: NO₂⁻, 0.1 mg L⁻¹; NO₃⁻, 0.05 mg L⁻¹; NH₄⁺, 1 mg L⁻¹. The method proposed has been successfully used for speciation analysis of inorganic nitrogen in precipitation.     

Two different spectrofluorimetric methods for simultaneous determination of gemfibrozil and rosiglitazone in human plasma

Two accurate, reliable, and highly sensitive spectrofluorimetric methods were developed for simultaneous determination of binary mixture gemfibrozil and rosiglitazone in human plasma without prior separation steps. The first method is based on synchronous fluorescence spectrometry using double scans. At Δλ = 27 nm, gemfibrozil yields detectable signal that is independent of the presence of rosiglitazone. Similarly, at Δλ = 120 nm the signal of rosiglitazone is not influenced by the presence of gemfibrozil. Signals at two wavelengths, 301 (Δλ = 27 nm) and 368 nm (Δλ = 120 nm) vary linearly with gemfibrozil and rosiglitazone concentrations over the range 100-700 ng mL⁻¹ (for gemfibrozil) and 20-140 ng mL⁻¹ (for rosiglitazone), respectively. The limits of detection (LOD) were 2.3 and 2.72 ng mL⁻¹ for gemfibrozil and rosiglitazone, respectively. The second method is based on the technique of simultaneous equations (Vierodt's method), in which 258 nm was selected as the excitation wavelength. Two equations are constructed based on the fact that at (λEm₂=302 nm of gemfibrozil) and (λEm₂=369 nm of rosiglitazone) the fluorescence of the mixture is the sum of the individual fluorescence of gemfibrozil and rosiglitazone. The limits of detection (LOD) were 28.1 and 23.63 ng mL⁻¹ for gemfibrozil and rosiglitazone, respectively. The proposed methods were successfully applied for the determination of the two compounds in synthetic mixtures and in human plasma with a good recovery.     

Fluorimetric determination of phytic acid in urine based on replacement reaction

A sensitive fluorimetric method for determination of phytic acid in human urine samples was described. The method was based on a fluorimetric replacement reaction, in which the added phytic acid replaced the Cu²⁺ ion from Cu²⁺-gelatin complex, liberating the fluorescent gelatin molecule. The fluorescence of the solution was accordingly recovered proportionally to the amount of the foreign phytic acid. The excitation wavelength was 273.5 nm and the characteristic emission wavelength was 305.0 nm, respectively. The calibration graph was obtained by plotting the recovered fluorescent intensity at maximum 305.0 nm against the added standard phytic acid, and was divided into two sections. One section was linear over the range of 0.40-2.40 mg L⁻¹ with a linear regression equation of I_f = −0.895 + 15.146c (R² > 0.9993), and the other over the range of 2.40-9.20 mg L⁻¹ with a linear regression equation of I_f = −29.526 + 26.113c (R² > 0.9996), respectively. The relative standard deviation (R.S.D.) at 95% confidence degree for a 2.0 mg L⁻¹ of standard phytic acid within 1 month was less than 1.26% (n = 5), indicating the procedure is reproducible. The detection and the quantification limits of phytic acid were estimated to be 0.23 and 0.40 mg L⁻¹, respectively. The proposed method was applied to the determination of phytic acid in urine samples and the found concentrations of phytic acid in urine were in the range of 0.49-0.75 mg L⁻¹ with recoveries of 96.2-108.8%. Comparison of the obtained results with the reported HPLC was performed, indicating the proposed method was reliable.