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%.     

Flow injection system with in-line Winkler's procedure using 16-way valve and spectrofluorimetric determination of dissolved oxygen in environmental waters

Flow injection spectrofluorimetry with in-line Winklers procedure was developed for the dissolved oxygen (DO) determination. 2-Thionaphthol reacted with iodine produced by Winkler’s method to form fluorescence inactive disulfide compound. To automate the process completely, a 5-channel flow system with a newly designed 16-way valve was assembled. The system consisted of a dispersion coil (DC), a precipitate formation coil (PFC), a precipitate dissolving coil (PDC), and extraction coil (EC). A calibration can be constructed by using a standard iodine solution for dissolved oxygen. The calibration graph was linear over the range 1.2×10⁻⁴∼6.0×10⁻⁴ mol l⁻¹ iodine (1.96∼9.80 mg O l⁻¹)). The relative standard deviation (n=6) was below 0.3% for the 4×10⁻⁴ mol l⁻¹ iodine (6.27 mg O l⁻¹) determination. The sample throughput was 12/h.     

Enzymatic chemiluminescent assay of glucose by sequential-injection analysis with soluble enzyme and on-line sample dilution

This work reports a sequential-injection analysis (SIA) method for the enzymatic assay of glucose with soluble glucose oxidase (GOD) and on-line sample dilution with chemiluminescence (CL) detection. A zone of sample was aspirated in the holding coil of the SIA manifold and, if necessary, was diluted on-line by means of an auxiliary dilution conduit. Then, a zone of GOD was aspirated adjacent to the sample zone and a stopped-flow period was applied to allow the enzymatic reaction to proceed with production of hydrogen peroxide. Then, zones of a catalyst (Co(II) solution) and alkaline luminol were aspirated into the holding coil. Finally, the flow was reversed and the stacked zones were sent to a flow-cell located in front of a photomultiplier tube (PMT) that monitored the CL intensity. The linear dynamic range was 1 × 10⁻⁵-1 × 10⁻³ mol L⁻¹ glucose, the coefficient of variation at 8 × 10⁻⁵ mol L⁻¹ of glucose was s_r = 3.1% (n = 8), the limit of detection at the 3σ level was c_L = 1 × 10⁻⁶ mol L⁻¹ and the sampling frequency was 28 h⁻¹. With on-line dilution by a factor of 1/200, the linear range could be extended up to 0.2 mol L⁻¹ glucose. The advantages of the proposed method are the simple manifold and instrumentation used, the scope for automated on-line dilution, the low consumption of sample and reagents and the elimination of enzyme immobilisation procedures. The method was applied to the analysis of commercial drinks and honey with percent relative errors in glucose determination in the range 100 ± 6.1%.     

Single interface flow analysis with accuracy assessment

Single interface flow systems (SIFA) present some noteworthy advantages when compared to other flow systems, such as a simpler configuration, a more straightforward operation and control and an undemanding optimisation routine. Moreover, the plain reaction zone establishment, which relies strictly on the mutual inter-dispersion of the adjoining solutions, could be exploited to set up multiple sequential reaction schemes providing supplementary information regarding the species under determination. In this context, strategies for accuracy assessment could be favourably implemented. To this end, the sample could be processed by two quasi-independent analytical methods and the final result would be calculated after considering the two different methods. Intrinsically more precise and accurate results would be then gathered.In order to demonstrate the feasibility of the approach, a SIFA system with spectrophotometric detection was designed for the determination of lansoprazole in pharmaceutical formulations. Two reaction interfaces with two distinct π-acceptors, chloranilic acid (CLA) and 2,3-dichloro-5,6-dicyano-p-benzoquinone (DDQ) were implemented.Linear working concentration ranges between 2.71 × 10^− 4 to 8.12 × 10^− 4 mol L^− 1 and 2.17 × 10^− 4 to 8.12 × 10^− 4 mol L^− 1 were obtained for DDQ and CLA methods, respectively. When compared with the results furnished by the reference procedure, the results showed relative deviations lower than 2.7%. Furthermore, the repeatability was good, with r.s.d. lower than 3.8% and 4.7% for DDQ and CLA methods, respectively. Determination rate was about 30 h^− 1.     

Sequential injection method for the direct spectrophotometric determination of bismuth in pharmaceutical products

A spectrophotometric method is reported for the determination of bismuth in pharmaceutical products using sequential injection analysis. Methylthymol blue (MTB) was used as a color forming reagent and the absorbance of the Bi(III)-MTB complex was monitored at 548 nm. The various chemical and physical variables that affected the reaction were studied. A linear calibration graph was obtained in the range 0.0-75.0 mg l⁻¹ Bi(III) at a sampling frequency of 72 h⁻¹. The reagent consumption was considerably reduced compared to conventional flow injection systems, as only 150 μl of MTB were consumed per run. The precision was very satisfactory (s_r=0.5%, at 50.0 mg l⁻¹ Bi(III), n=12) and the limit of detection, c_L, was 0.250 mg l⁻¹. The developed method was applied successfully to the analysis of various pharmaceutical products containing Bi(III). The relative errors e_r, were <1.5% in all cases and were evaluated by comparison of the obtained results with those found using atomic absorption spectrometry as the reference method.     

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.     

Spectrophotometric flow-injection determination of copper and nickel in plant digests exploiting differential kinetic analysis and multi-site detection

A novel strategy for implementing differential reaction-rate methods in flow-injection analysis is proposed and applied to the determination of copper and nickel in plant digests using 2-(5-brom-2-pyridylazo)-5-(diethylamino)-phenol (Br-PADAP) as the color-forming reagent. Multi-site detection is involved, therefore the flow cell is displaced between two monitoring sites, and the analytical signals refer to different conditions of sample dispersion, reaction development and timing.The system handles 20 samples h⁻¹ and requires 0.32 mg Br-PADAP per determination. Signal additivity was evaluated within 98 and 102%, and linear responses (r > 0.999; n = 6) were verified for both copper and nickel up to 0.80 mg l⁻¹. Detection limits of 0.01 and 0.04 mg l⁻¹ Cu and Ni were estimated by considering the highest concentration of the counter analyte. Results are precise (R.S.D. < 2%) and in agreement with ICP-OES (95% confidence level). Potentialities and limitations of the approach are discussed.     

Radial basis network analysis of color parameters to estimate lycopene content on tomato fruits

With the purpose of estimating the lycopene concentration in tomato food samples, in an non-destructive way, several types of linear models of color parameters have been tested using individual values of L*, a* and b* values, (a*/b*), (a * ²/b * ²) and chroma parameters from tomato juice and fresh tomato fruits obtained with two different apparatus (Minolta CR-200b triestimulus colorimeter and HunterLab LabScan XE). Lycopene concentrations of fresh tomato and tomato juice (used as an input) were analyzed by UV-Vis spectroscopy. For all linear methods applied, the best one to estimate the lycopene concentration in tomato was the L*, a* and b* values of tomato juice measured with Hunter colorimeters (adjusted correlation coefficient, and mean prediction error, MPE < 6.59%). Four different RBEF models were designed firstly using three color parameters (L*, a* and b*) designated as “Lab case”, and secondly individually by the (a*/b*), (a * ²/b * ²) and chroma parameters. The lycopene concentration estimations were carried out with the lowest MPE and highest values possible. In order to test the reliability of the non-linear models, external validation process was also performed. From the testing of the all non-linear models applied, the RBEF Lab case model was the best to estimate lycopene content from color parameters (L*, a* and b*) using Minolta or Hunter equipments (MPE lower than 0.009 and higher than 0.997). This was a simple non-destructive method for predicting lycopene concentration in tomato fruits and tomato juice, which was reproducible and accurate enough to substitute chemical extraction determinations, and may be a useful tool for tomato industry.     

Flow injection wetting-film extraction system for flame atomic absorption spectrometric determination of cadmium in environmental waters

A newly simple flow injection wetting-film extraction system coupled to flame atomic absorption spectrometry (FAAS) has been developed for trace amount of cadmium determination. The sample was mixed on-line with sodium diethyl dithiocarbamate and the produced non-charged Cd(II)-diethyl dithiocarbamate (DDTC) chelate complex was extracted on the thin film of diisobutyl ketone (DIBK) on the inner wall of the PTFE extraction coil. The wetting-film with the extracted analyte was then eluted by a segment of the cover solvent, and transported directly to the FAAS for evaluation. All the important chemical and flow parameters were optimized. Under the optimized conditions an enhancement factor of 35, a sample frequency of 22 h⁻¹ and a detection limit of c_L = 0.7 μg l⁻¹ Cd(II) were obtained for 60 s preconcentration time. The calibration curve was linear over the concentration range 1.5-45.0 μg l⁻¹ Cd(II) and the relative standard deviation, R.S.D. (n = 10) was 3.9%, at 10.0 μg l⁻¹ concentration level. The developed method was successfully applied to cadmium determination in a variety of environmental water samples as well as waste-water sample.     

Characterization and evaluation of a macroporous adsorbent for possible use in the expanded bed adsorption of flavonoids from Ginkgo biloba L.

The suitability of the use of macroporous adsorbent Amberlite XAD7HP in expanded bed adsorption processes for the isolation of flavonoids from crude extracts of Ginkgo biloba L. has been assessed. The expansion and hydrodynamic properties of expanded beds were investigated and analyzed. The bed expansion as a function of operational fluid velocity was measured and correlated with the Richardson–Zaki equation. Theoretical predictions of the correlation parameters (the terminal settling velocity u_t and exponent n) were improved by modifying equations in the literature. Residence time distributions (RTDs) were studied using acetone as a tracer. Three measures of liquid phase dispersion (the height equivalent of theoretical plate, Bodenstein number and axial distribution coefficient) were investigated and compared to values previously obtained with commercial EBA adsorbents developed for protein purification. A suitable bed expansion ratio was found to be 1.25 times the settled bed height, which occurred at a corresponding flow velocity of 183 cm/h. For an initial settled bed height of 42 cm, the mean residence time of liquid in the expanded bed was around 28 min. Under these flow conditions, the axial mixing coefficient D_ax was 7.54 × 10⁻⁶ m²/s and the Bodenstein number was 28; the number of theoretical plates (N) was 19 and the height equivalent of a theoretical plate (HETP) was 2.77 cm. Rutin trihydrate was used as a model flavonoid for the characterization of the adsorption properties of Amberlite XAD7HP. Adsorption was observed to reach equilibrium within 3 h with 70% of the adsorption capacity being achieved within 30 min. The estimated maximum equilibrium adsorption capacity for rutin was estimated to be 43.0 mg/(g resin) when the results were fitted to Langmuir isotherms. The adsorption performance was not seriously impaired by the physical presence of G. biloba leaf powders. Assessment of the kinetics of the adsorption of rutin revealed that the rate constant for adsorption was only reduced by 15% in the presence of leaf powders at a concentration of 50 mg/mL. The results demonstrated that Amberlite XAD7HP should be suitable for expanded bed adsorption of flavonoids from crude extracts of G. biloba L.     

Zwitterionic hydrophilic interaction chromatography coupled with post-column derivatization for the analysis of glutathione in wine samples

In this study the development, validation and application of a new chromatographic method for the determination of glutathione (GSH) in wine samples is presented. The separation of the GSH was carried out using a sulfobetaine-based hydrophilic interaction chromatography (HILIC) analytical column whereas its detection was carried out spectrofluorimetrically (λ_ext/λ_em = 340/455 nm) after post-column derivatization with o-phthalaldehyde. GSH was separated efficiently from matrix endogenous compounds of wines by using a mobile phase of 15 mmol L⁻¹ CH₃COONH₄ (pH = 2.5)/CH₃CN, 35/65% (v/v). The parameters of the post-column reaction (pH, amount concentration of the reagent and buffer solution, flow rate, length of the reaction coil) were investigated. The linear determination range for GSH was 0.25–5.0 μmol L⁻¹ and the LOD was 19 nmol L⁻¹. No matrix effect was observed, while the accuracy was evaluated with recovery experiments and was ranged between 89% and 108%.     

A sequential injection system for indirect spectrophotometric determination of lactic acid in yogurt and fermented mash samples

A suitable non-enzymatic method is presented as an alternative to the lactic acid determination in yogurt and fermented mash samples. The oxidative conversion of lactic acid by Ce⁴⁺ to CO₂ was performed in a sequential injection system with a heating coil set at 45 °C. A gas diffusion unit was coupled to the flow system for promoting the permeation of CO₂, which was collected into a bromothymol blue solution (pH 8.4), used as indicator solution for the spectrophotometric determination (619 nm). Simplicity in operation, low reagent consumption, low cost and ruggedness are some remarkable characteristics of the proposed system. Base line drift was < 0.005 h^− 1. A linear range from 20.0 to 100.0 mg L^− 1 lactic acid was obtained (r² = 0.998), and the detection and quantification limits were estimated as 0.158 mg L^− 1 and 1.6 mg L^− 1, respectively. The sampling rate was 22 h^− 1, with a consumption of 0.04 g Ce⁴⁺ per determination. Interferences of matrix components were not detected. Samples of yogurt and sugar cane fermented mash products were analyzed, and no significant difference at 95% confidence level was observed when comparing the proposed method with HPLC analysis.     

Absorbance characteristics of a liquid-phase gas sensor based on gas-permeable liquid core waveguides

The absorbance characteristics and influential factors on these characteristics for a liquid-phase gas sensor, which is based on gas–permeable liquid core waveguides (LCWs), are studied from theoretical and experimental viewpoints in this paper. According to theory, it is predicted that absorbance is proportional to the analyte concentration, sampling time, analyte diffusion coefficient, and geometric factor of this device when the depletion layer of the analyte is ignored. The experimental results are in agreement with the theoretical hypothesis. According to the experimental results, absorbance is time-dependent and increasing linearly over time after the requisite response time with a linear correlation coefficient r² > 0.999. In the linear region, the rate of absorbance change (RAC) indicates improved linearity with sample concentration and a relative higher sensitivity than instantaneous absorbance does. By using a core liquid that is more affinitive to the analyte, reducing wall thickness and the inner diameter of the tubing, or increasing sample flow rate limitedly, the response time can be decreased and the sensitivity can be increased. However, increasing the LCW length can only enhance sensitivity and has no effect on response time. For liquid phase detection, there is a maximum flow rate, and the absorbance will decrease beyond the stated limit. Under experimental conditions, hexane as the LCW core solvent, a tubing wall thickness of 0.1 mm, a length of 10 cm, and a flow rate of 12 mL min⁻¹, the detection results for the aqueous benzene sample demonstrate a response time of 4 min. Additionally, the standard curve for the RAC versus concentration is RAC = 0.0267 c + 0.0351 (AU min⁻¹), with r² = 0.9922 within concentrations of 0.5–3.0 mg L⁻¹. The relative error for 0.5 mg L⁻¹ benzene (n = 6) is 7.4 ± 3.7%, and the LOD is 0.04 mg L⁻¹. This research can provide theoretical and practical guides for liquid–phase gas sensor design and development based on a gas-permeable Teflon AF 2400 LCW.     

Studies on photocrosslinkable copolymers of 4-methacryloyloxyphenyl-3′,4′-dimethoxystyryl ketone and methyl methacrylate

Copolymers with various contents of 4-methacryloyloxyphenyl-3′,4′-dimethoxystyryl ketone (MPDSK) and methyl methacrylate (MMA) were prepared in methyl ethyl ketone solution using benzoyl peroxide as a free radical initiator at 70 °C. Characterization of the resulting polymers was done by UV, FT-IR, ¹H NMR and ¹³C NMR spectroscopic techniques. The copolymer compositions were determined by ¹H NMR analysis. The monomer reactivity ratios were calculated using linearisation methods such as Finemann-Ross (r₁ = 0.4283 and r₂ = 0.3050), Kelen-Tudos (r₁ = 0.4264 and r₂ = 0.2606), and extended Kelen-Tudos (r₁ = 0.4022 and r₂ = 0.2704) methods as well as by a non-linear error-in-variables model (EVM) method using the computer program RREVM (r₁ = 0.4066 and r₂ = 0.2802). The molecular weights ( and ) and the polydispersity index of the copolymers were determined by gel permeation chromatography. The thermal stability of the copolymers increases with increase in concentration of MPDSK. Glass transition temperatures were determined by differential scanning calorimeter under nitrogen atmosphere. The photoreactivity of the copolymers having pendant chalcone moieties was studied in chloroform solution.     

Liquid-liquid extraction procedure exploiting multicommutation in flow system for the determination of molybdenum in plants

A liquid-liquid extraction flow analysis procedure for the spectrophotometric determination of molybdenum in plants at μg l⁻¹ level is described. The flow network comprised a set of solenoid valves assembled to implement the multicommutation approach under microcomputer control. Radiation source (LED, 475 nm), detector (photodiode) and separation chamber were nested together with the flow cell comprising a compact unit. The consumption of reagents (potassium thiocyanate and stannous chloride) and also extracting solvent (isoamyl alcohol) were optimized to 32 mg and 200 μl per determination, respectively. Accuracy was assessed by comparing results with those obtained with ICP-OES and no significant difference at 95% confidence level was observed. Other favorable characteristics such as a linear response ranging from 25 to 150 μg l⁻¹ molybdenum (r=0.999); detection limit of 4.6 μg l⁻¹ sample throughput of 25 determinations per hour and relative standard deviation of 2.5% (n=10) were also achieved.     

Synthesis and characterisation of molecularly imprinted catalytic microgels for carbonate hydrolysis

Novel molecularly imprinted microgels incorporating arginine and tyrosine side chains as functional monomers have been designed and synthesised with percentages of cross-linker ranging from 70 to 90%. Full chemico-physical characterisation including M_r, coil density and size particle determination concluded that all polymer preparations obtained can be classified as microgels. Molecular imprinting using a phosphate template was used to generate catalytic microgels for the hydrolysis of p-nitrophenyl carbonates. Kinetic characterisation of the catalytic activity of the different preparations indicated that values of critical monomer concentration (C_M) and percentage of cross-linker play an important role in determining the catalytic efficiency of the different preparations. Microgels containing 70% cross-linker were the only ones following the Michaelis-Menten saturation model and kinetic parameters were obtained using 4 mg/ml of M397: V_max = 1.34 × 10⁻⁶ M s⁻¹ (S.E. 1.28 × 10⁻⁷) and K_M = 2.38 × 10⁻³ M (S.E. 3.1 × 10⁻⁴).     

A compact portable flow analysis system for the rapid determination of total phosphorus in estuarine and marine waters

The development and evaluation of a portable flow analysis system for the in situ determination of total phosphorus is described. The system has been designed with rapid underway monitoring in mind. The system employs an ultra-violet photo-reactor and thermal heating for peroxodisulfate digestion of total phosphorus to orthophosphate, followed by spectrophotometric detection with a multi-reflective flow cell and low-power light emitting diode using the molybdenum blue method. Reagents are stored under gas pressure and delivered using software controlled miniature solenoid valves. The fully automated system has a throughput of 115 measurements per hour, a detection limit of 1 μg P L⁻¹, and gives a linear response over the calibration range of 0-200 μg P L⁻¹ (r² = 0.9998), with a precision of 4.6% RSD at 100 μg P L⁻¹ (n = 10). Field validation of the instrument and method was performed in Port Philip and Western Port Bays in Victoria, SE Australia, where 2499 analyses were performed over a 25 h period, over a cruise path of 285 km. Good agreement was observed between determinations of samples taken manually and analysed in the laboratory and those measured in situ with the flow analysis system.     

Amperometric determination of chloroguaiacol at submicromolar levels after on-line preconcentration with molecularly imprinted polymers

In this study, a sorbent flow preconcentration system coupled to amperometric detector for the chloroguaiacol (4-chloro-2-methoxyphenol) determination at submicromolar levels is described. The satisfactory selectivity of the proposed method was attained by means of the use of a chloroguaiacol-imprinted polymer, whose the synthesis was carried out by bulk polymerization. Flow and chemical parameters associated to the preconcentration system, such as sample pH, preconcentration and elution flow rates, concentration of the carrier solution (KCl) and eluent volume were investigated through multivariate analysis. The flow preconcentration of chloroguaiacol was not affect by equimolar presence of structurally similar phenolic compounds including catechol, 4-chloro-3-methylphenol, 4-aminophenol and 2-cresol, thus showing the good performance of the imprinted polymer. Under the best experimental conditions, it was obtained a preconcentration factor of 110-fold and low detection and quantification limits of 27 and 78 nmol L⁻¹, respectively. The analytical curve covered a wide linear range from 0.05 up to 5.0 μmol L⁻¹ (r > 0.999) and satisfactory precision (n = 8) evaluated by relative standard deviation (R.S.D.) were respectively, 5.5 and 4.2%, for solutions of 1.0 and 5.0 μmol L⁻¹ chloroguaiacol. Other parameters related to the performance of the flow system were also evaluated including concentration efficiency of 27.5 min⁻¹ and consumptive index of 0.09 mL. Recoveries varying from 93 up to 112% for water samples (tap water and river water) spiked with chloroguaiacol concentration were achieved, thus assuring the accuracy of the proposed flow preconcentration system.     

A very sensitive flow system for the direct determination of copper in natural waters based on spectrophotometric detection

A very sensitive flow injection method with spectrophotometric detection has been developed for the on-line determination of copper in natural waters. The method exhibits a limit of detection three times lower than the most sensitive direct spectrophotometric method previously described and then allows the direct and simple in situ determination of copper in most natural waters.The method was based on the measurement of the absorbance of the coloured complex formed by copper with the chromogenic reagent di-2-pyridyl ketone benzoylhydrazone (dPKBH) in an alkaline medium. This complex presents stoichiometry 1:2 (Cu:dPKBH), and exhibits maximum absorbance at 370 nm. The manifold used was very simple, and consisted of two channels. The first one contained the sample while the second one contained the colorimetric reagent (3.3×10⁻⁴ M dPKBH in 10% ethanol), in a 1.6×10⁻² M phosphate buffer solution at pH 8. The performance of the system was optimised by using both univariate and modified simplex methodologies. When modified simplex was used, the best signal was obtained for a sample injection volume of 529 μl, a reaction coil length of 1.29 m, and a reagent flow rate of 4.8 ml min⁻¹. Under optimum conditions, the response was linear up to 3 mg l⁻¹ copper, the equation of the straight line being y=0.314x+5.2×10⁻⁴ (r²=0.998). The method allowed a sampling frequency of 40 samples per hour and exhibited a precision of 2.11% (as R.S.D., n=11). The limit of detection was 4.6 μg l⁻¹ (calculated as 3s_b/m, where s_b is the standard deviation of the y-intercept and m represents the slope of the straight line), and was therefore more sensitive than all the direct continuous methods reported previously.The method was successfully applied to the analysis of real water samples, with an average relative error of 5.32%.