Simultaneous determination of saccharin and aspartame in commercial noncaloric sweeteners using the PLS-2 multivariate calibration method and validation by capillary electrophoresis

A new method to determine a mixture for sweetener sodium saccharin and aspartame in commercial noncaloric sweeteners is proposed. A classical full factorial design for standards was used in the calibration step to build the partial least-squares (PLS-2) model. Instrumental data were obtained by means of UV-visible spectrophotometry. Salicylic acid was used as an internal standard to evaluate the adjustment of the real samples to the PLS model. The concentration of analytes in the commercial samples was evaluated using the obtained model by UV spectral data. The PLS-2 method was validated by capillary zone electrophoresis (CZE), finding in all cases a relative error of less than 11% between the PLS-2 and the CZE methods. The proposed procedure was applied successfully to the determination of saccharin and aspartame in noncaloric commercial sweeteners.     

Determination of seven artificial sweeteners in diet food preparations by reverse-phase liquid chromatography with absorbance detection

The artificial sweeteners aspartame, saccharin, cyclamate, alitame, acesulfam-K, sucralose, and dulcin are determined in diet soft drinks and tabletop sweetener preparations. Samples are diluted, filtered, and analyzed directly by liquid chromatography on a C-18 reverse-phase column with a mobile phase gradient ranging from 3% acetonitrile in 0.02M KH2PO4 (pH 5) to 20% acetonitrile in 0.02M KH2PO4 (pH 3.5). Diet puddings and dessert toppings are extracted with ethanol, filtered, and diluted with mobile phase for analysis. The sweeteners, except sucralose and cyclamate, were detected by UV absorbance at either 200 or 210 nm. Sucralose was determined at 200 nm or by refractive index. Cyclamate was determined after post-column ion-pair extraction. The sweeteners stevioside and talin were not detected. Additives such as caffeine, sorbic acid, and benzoic acid did not interfere.     

Orthophosphate, phytate, and total phosphorus determination in cereals by flow injection analysis

A flow injection spectrophotometric procedure with enzymatic hydrolysis was developed for determination of orthophosphate, phytate and total phosphorus in cereal samples. Phosphorus species were extracted from cereals with 0.05 mol L(-1) potassium hydrogen phthalate buffer solution at pH 5.7. Orthophosphate was directly determined in the extracts by molybdenum blue spectrophotometric method. The phytate was hydrolyzed by the enzyme phytase coupled to a solid phase packed into an enzymatic reactor, and the resulting hydrolyzed orthophosphate was also determined by spectrophotometry at 650 nm. After optimization for phosphorus species extraction and enzymatic hydrolysis, a linear calibration graph was obtained up to 196 x 10(-6) mol L(-1) orthophosphate (P conc = -2.67 + 0.52x, r = 0.9998). Measurements are characterized by relative standard deviation of 1.6% for a standard of 72 x 10(-6) mol L(-1) orthophosphate and no baseline drift was observed during 4 h operation periods. It provides 72 measurements per hour, with 2.4 x 10(-)6) mol L(-1) and 7.9 x 10(-6) mol L(-1) as detection and quantification limits, respectively.     

Flow injection potentiometric system for the simultaneous determination of inositol phosphates and phosphate: phosphorus nutritional evaluation on seeds and grains

A simple flow injection potentiometric (FIP) system, which uses a tubular cobalt electrode, has been developed for phosphorus nutritional evaluation of seeds and grains. Inorganic phosphorus, P(i), is determined using a 1 x 10(-2) mol.L(-1) potassium phthalate buffer solution adjusted at pH 4. A sensitivity of 47 mV/decade and an operating range from 10 to 1000 mg.L(-1) (1 x 10(-4)-1 x 10(-2) M) of dihydrogen phosphate are obtained. The inositol phosphates amount, which is referred to the organic phosphorus, P(org), is directly determined from extracts using a 1 x 10(-2) mol.L(-1) Tris-HCl buffer solution adjusted at pH 8. A sensitivity of 127 mV/decade and an operating range of 10-1000 mg.L(-1) (2.5 x 10(-4)-5 x 10(-3) M) of P(org) (expressed as inositol hexakisphosphoric acid monocalcium) are achieved. Some samples of seed and grain are analyzed by an ICP-OES and a spectrophotometric method to compare results to the developed flow system; no significant differences at the 95% confidence level are observed using a paired t test. Other samples such as animal nursing feed, soybean meal, and corn are also analyzed with the proposed FIP system, showing a good correlation to the ICP-OES values.     

Spectrophotometric determination of isocarboxazid bulk drug and tablets by reaction with p-dimethylaminocinnamaldehyde

A spectrophotometric method is described for the determination of isocarboxazid. The method is based on the reaction of the drug with p-dimethylaminocinnamaldehyde in the presence of trichloroacetic acid in a methanolic medium to produce a very intense red chromophore (lambda max = 500 nm, Emax = 1.05 x 10(5]. The reaction is proposed to proceed via electrophilic attack at the C-4 position of the isoxazole nucleus. Job's plot indicated a 1:1 drug-to-reagent ratio. Regression analysis of Beer's plot showed excellent correlation (r = 0.9996) in the concentration range 0.25-2.10 micrograms isocarboxazid/mL. The developed color is stable for at least 12 h. Results of analyses of bulk drug and tablets by the proposed method are comparable to those for USP XXI methods.     

Study on the supramolecular interaction of thiabendazole and beta-cyclodextrin by spectrophotometry and its analytical application

The beta-cyclodextrin-thiabendazole (beta-CD-TBZ) inclusion complex was synthesized and its structure characterized by (1)H NMR and IR. The mechanism of the supramolecular interaction of TBZ and beta-CD has been studied and discussed by spectrophotometry. The results showed that the phenyl ring of TBZ was included in the beta-CD cavity to form a 1:1 host-guest complex with an apparent formation constant of 1.60 x 10(3) mol(-1).L. On the basis of the enhancement of the absorbance of TBZ produced through complex formation, a spectrophotometric method for the determination of TBZ in bulk aqueous solution in the presence of beta-CD was developed. The linear relationship between the absorbance and TBZ concentration was obtained in the range of 8.86 x 10(-7)-1.45 x 10(-5) mol/L. The detection limit was 2.71 x 10(-7)mol/L, and the relative standard deviation was 0.86%. The interference of 48 coexisting substances was slight. The proposed method has been successfully applied to the determination of TBZ in fruits with recoveries of 96-103%.     

Isolation and characterization of aminopeptidase (Jc-peptidase) from Japanese cedar pollen (Cryptomeria japonica)

An aminopeptidase, Jc-peptidase, was purified from Japanese cedar pollen by seven steps, including precipitation with ammonium sulfate, ion-exchange chromatography, gel filtration, hydrophobic interaction chromatography on phenyl-agarose, and high-performance liquid chromatography. Purified Jc-peptidease has a molecular weight of 42 kDa and hydrolyzes the synthetic substrates of L-phenylalanyl-4-methylcoumaryl-7-amide (Phe-MCA) with Km = 5 x 10(-5) M, Tyr-MCA with Km = 7 x 10(-4) M, Leu-MCA with Km = 1 x 10(-3) M, and Met-MCA with Km = 1 x 10(-3) M. Other MCA analogues such as Arg-MCA or Glu-MCA failed to serve as its substrates. The activity was inhibited in the presence of phebestin, [(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-L-valyl]-L-phenylalanine, with Ki = 4.7 x 10(-5) M, or bestatin, [(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl]-L-leucine, with Ki = 1.1 x 10(-4) M. According to amino acid sequence analysis, the N-terminal amino group seems to be blocked. The physiological function of the aminopeptidase (Jc-peptidase) has not been clarified in vivo.     

Colorimetric determination of hydralazine with 9-chloroacridine

A spectrophotometric assay for hydralazine hydrochloride based on the interaction of the drug with 9-chloroacridine has been developed. The interaction shows an absorption maximum at 460 nm and is affected by temperature, heating time, and quantity of acridine reagent used. Color development is maximum when the drug is heated in the presence of a 30-fold molar excess of the acridine in a 50 +/- 1 degree C water bath for 1 hr. The method detects hydralazine hydrochloride in the 10(-5)--10(-6)M range with sensitivity to 0.2 micrograms/mL and 3--4% accuracy. Typical calibration data obtained from linear regression analysis of absorbance at various drug concentrations show r = 0.9997 (n = 6). Hydralazine can be determined in dosage forms that also contain varying quantities of reserpine and hydrochlorothiazide. The 9-chloroacridine method is as sensitive as other spectrophotometric procedures for hydralazine but also more accurate and precise, and involves fewer manipulative steps.     

Metabolism of stevioside and rebaudioside A from Stevia rebaudiana extracts by human microflora

Stevia rebaudiana standardized extracts (SSEs) are used as natural sweeteners or dietary supplements in different countries for their content of stevioside or rebaudioside A. These compounds possess up to 250 times the sweetness intensity of sucrose, and they are noncaloric and noncariogenic sweeteners. The aim of this study was to investigate the in vitro transformation of stevioside and rebaudioside A after incubation with human microflora, the influence of these sweeteners on human microbial fecal community and which specific groups metabolize preferentially stevioside and rebaudioside A. The experiments were carried out under strict anaerobic conditions in batch cultures inoculated with mixed fecal bacteria from volunteers. The hydrolysis was monitored by HPLC coupled to photodiode array and mass spectrometric detectors. Isolated bacterial strains from fecal materials incubated in selective broths were added to stevioside and rebaudioside A. These sweeteners were completely hydrolyzed to their aglycon steviol in 10 and 24 h, respectively. Interestingly, the human intestinal microflora was not able to degrade steviol. Furthermore, stevioside and rebaudioside A did not significantly influence the composition of fecal cultures; among the selected intestinal groups, bacteroides were the most efficient in hydrolyzing Stevia sweeteners to steviol.     

Determination of the hydroperoxil group level in polyethylene glycols using a novel mimic enzyme Fe(III)-Mn(II)(HNAPTS)2

Two kinds of Schiff base metal complexes of 2-hydroxy-1-naphthaldehyde-2-amino-5-phenylthiazole (HANPTS), Mn(II)(HNAPTS)(2) and Fe(III)(HNAPTS)(2), were synthesized and used to mimic the active group of horseradish peroxidase (HRP). The catalytic characteristics of the mimic enzymes in the oxidation reaction of ascorbic acid (AsA) with the OOH group in polyethylene glycols (PEGs) have been studied by a spectrophotometric method. Fe(III) has remarkable coordinated catalysis to Mn(II)(HNAPTS)(2); as a result, the catalytic ability of Fe(III)-Mn(II)(HNAPTS)(2) is 75% of that of HRP. The possible mechanism of the reaction was discussed. The linear relationship between deltaA(265)(AsA) and OOH group concentrations was in the range of 1.5 x 10(-6) to 9.0 x 10(-4) mol/L. The proposed method was successfully applied to the determination of the OOH group level in different molecular weight PEGs.     

Enzymatic determination of urea in milk by sequential injection with spectrophotometric and conductometric detection

In this work, an analytical system based on the coupling of gas diffusion separation and sequential injection analysis for urea determination in milk is presented. A versatile manifold that could simultaneously be used for either spectrophotometric or conductometric detection was constructed. The sample and urease solution are sequentially aspirated into the holding coil and sent to a thermoreactor, where urea is enzymatically hydrolyzed by urease and converted into ammonium. This stream merges an alkaline solution at a confluence point where ammonia is formed. Ammonia diffuses through a hydrophobic membrane and modifies the bromothymol blue indicator color, when spectrophotometric detection is used, or changes the conductance of a boric acid solution acceptor stream, when conductometric detection is used.This methodology was applied to the determination of urea in 18 milk samples and the results were statistically comparable with those furnished by the enzymatic recommended procedure. The detection limits were 2.6 x 10(-4) and 2.8 x 10(-5) mol L(-1) for conductometric and spectrophotometric detection, respectively. Repeatability (relative standard deviation, RSD) was better than 3.7% and 2.6% for conductometric and spectrophotometric detection, respectively.     

Reverse-phase liquid chromatographic determination of benzoic and sorbic acids in foods

An isocratic liquid chromatographic (LC) technique is described for the determination of benzoic acid and sorbic acid in foods such as beverages, fruits, seafood, vegetables, sauces, and dairy, bakery, and confectionery products. A C18 column is used with methanol-phosphate buffer (5 + 95) as mobile phase and 4-hydroxyacetanilide or 3,5-dinitrobenzoic acid as internal standard. Sample preparation is simple, rapid, and produces a sample extract that has a minimum effect on the column performance and life. Specificity of the method was checked against common food additives such as L-ascorbic acid, caffeine, artificial sweeteners (saccharin, cyclamate, aspartame), antioxidants (BHT, BHA) and artificial colors. Also described are 2 procedures for confirmation of the preservatives, using either redox reaction of sorbic acid with potassium permanganate or gas chromatography/mass spectrometry. Mean recoveries of 90-105% were obtained with a precision of 1-6% and a detection limit of 20 mg/kg for the 2 preservatives.     

Ultraviolet spectrophotometric determination of hydralazine hydrochloride in tablets following derivatization with nitrite

Routine use of the USP XXI spectrophotometric method for the content uniformity determination of hydralazine hydrochloride tablets has shown that tablet excipients can significantly alter the spectral characteristics of the drug and thus cause inaccurate assay values to be obtained. Because of this problem, a simple and reliable alternative spectrophotometric assay method, based on the conversion of hydralazine to tetrazolo [5,1-alpha]phthalazine with nitrite ions under acidic conditions, was developed. The derivative showed an absorption maximum at about 274 nm and obeyed Beer's law over the concentration range 4-40 micrograms/mL. Mean recoveries of hydralazine hydrochloride added to commercial coated and uncoated tablets were 101.0% (n = 10) and 100.8% (n = 8), respectively. The proposed method was found suitable for the assay not only of individual tablets but also of tablet composites.     

Accurate determination of oosporein in fungal culture broth by differential pulse polarography

A simple and accurate differential pulse polarographic method has been developed for the determination of oosporein in the culture broth of the fungus Beauveria brongniartii. This hydroxybenzoquinone derivative is the only major secondary metabolite secreted by this entomopathogenic fungus, which is used as biological pest control agent (BCA) against Melolontha melolontha larvae. It can be found in the host organism as well as in the formulated product. The polarographic behavior of oosporein was examined in various buffer systems over the pH range 3-10. In Britton-Robinson buffer/methanol solution (3:7 v/v, pH 5.5) the differential pulse polarograms exhibited reproducible peaks at E(p) = -0.18 V vs silver/silver chloride/potassium chloride (3 M). Under these conditions, a plot of peak height vs concentration of oosporein was found to be linear over the range 5.9 x 10(-)(7) to 2.5 x 10(-)(5) M (0.18-7.74 microg mL(-)(1); r = 0.9998). The detection limit was calculated to be 54 ng mL(-)(1). To evaluate the concentration of oosporein, the standard addition method was applied. The analysis of oosporein in the culture broth led to a mean value of 524.9 microg mL(-)(1) broth with a relative standard deviation (S(rel)) of +/-2.6%. The proposed polarographic method is accurate, not time-consuming, and it is of low cost because no separation steps are necessary.     

Improved water solubility of neohesperidin dihydrochalcone in sweetener blends

Significant technological advantages in terms of sweetness synergy and hence cost-saving can be obtained if neohesperidin dihydrochalcone (NHDC) is used in sweetener blends with other intense or bulk sweeteners. The combination of NHDC with sodium saccharin or sodium cyclamate is an excellent method to improve the water solubility at room temperature of NHDC. In the case of NHDC-sodium saccharin, two different methods for blend preparation, a simple mixture and a cosolubilized mixture, can be used, with similar results obtained for solubility and solution stability properties. To improve temporally the water solubility of NHDC in combination with sodium cyclamate, it is absolutely necessary to prepare cosolubilized blends.     

Liquid chromatographic determination of quinine, hydroquinine, saccharin, and sodium benzoate in quinine beverages

A liquid chromatographic (LC) method is described for the determination of quinine, hydroquinine, sodium saccharin, and sodium benzoate in soft drinks. The method involves simple sample preparation, direct injection onto an octadecylsilane column, and elution with a methanol-acetonitrile-water-acetic acid (20 + 10 + 70 + 1) mobile phase. Eluted constituents are measured spectrophotometrically at 254 nm. The relationship between peak height and concentration was linear between 20 and 120 micrograms/mL for quinine. A relative standard deviation of 0.82% was obtained for commercial samples spiked with quinine, and the average recovery was 100.3%. The proposed procedure is accurate and rapid and can also detect hydroquinine (a natural contaminant of quinine), sodium saccharin, and sodium benzoate. Linear responses ranged from 0.45 to 20 micrograms/mL for hydroquinine, from 54.8 to 219 micrograms/mL for sodium saccharin, and from 10.1 to 145.1 micrograms/mL for sodium benzoate. The reproducibility of the LC method was evaluated with standard solutions of hydroquinine, sodium saccharin, and sodium benzoate, which produced relative standard deviations of 0.42, 0.46, and 1.13%, respectively. The average recoveries for sodium saccharin and sodium benzoate from spiked samples were 99.4 and 100.2%, respectively.     

Preservatives and artifical sweeteners.

Twenty saccharin-containing table sweeteners, in the form of tablets, liquids, crystals, or blends, and manufactured or distributed by 15 different companies, were analyzed for their o-toluenesulfonamide (o-TS) content. A previously described procedure for the determination of o-TS in commercial saccharin was found to be applicable to the determination of o-TS in these preparations. o-TS was found in all analyzed samples, in amounts ranging from 57 to 3811 ppm. In some cases the concentration of o-TS varied even from lot to lot from the same manufacturer. One pair of lots contained 57 and 67 ppm o-TS, respectively, while in another 711 and 3003 PPM O-TS were found. Gas-liquid chromatographic patterns of samples from the same distributor (or manufacturer) were similar and characteristic of the brand. Recoveries of o-TS from liquid and tablet preparations were almost quantitative, while recoveries from saccharin blends were in the 95-103% range.     

Development of an extractive spectrophotometric method for the determination of copper(II) in leafy vegetable and pharmaceutical samples using pyridoxal-4-phenyl-3-thiosemicarbazone (PPT)

A highly sensitive extractive spectrophotometric method has been developed for the determination of copper(II) using pyridoxal-4-phenyl-3-thiosemicarbazone(PPT) as an analytical reagent. The PPT forms reddish brown species of copper(II) at a pH range of 3.0-5.5, and the complex was extracted into n-butanol. The Cu(II)-PPT complex shows maximum absorbance at 440 nm, with molar absorptivity and Sandell's sensitivity being 2.16 x 10(4) L mol(-1) cm(-1) and 2.94 x 10(-3) microg cm(-2), respectively. The system obeys Beer's law in the range of 0.2-5.0 mg/L. The regression coefficient of the Beer's law straight line is 0.338, and the correlation coefficient is 0.96. The detection limit of the method is 0.0065 microg mL(-1). Most of the common metal ions generally found associated with copper do not interfere. The repeatability of the method was checked by finding the relative standard deviation. The developed method has been successfully employed for the determination of copper(II) in leafy vegetable and pharmaceutical samples. The method is evaluated by analyzing samples from the Bureau of Analyzed Samples (BCS 233, 266, 216/1, 207, and 179) and by intercomparison of experimental values using AAS.     

Determination of trace hydroxyl radicals by flow injection spectrofluorometry and its analytical application

On the basis of the fluorescence increase of the reaction of ninhydrin with hydroxyl radicals, a new method for the determination of trace amounts of hydroxyl radicals by flow injection spectrofluorometry is presented. The introduction of flow injection analysis brought better reproducibility and avoided the effect of oxygen and other substances in the environment on the reaction of ninhydrin with hydroxyl radicals. Under optimum experimental conditions, the hydroxylated product of ninhydrin had excitation and emission maxima at 300 and 406 nm, respectively. The linear range was 2.60 x 10(-7) to 4.00 x 10(-5) M, and the limit of detection was 7.91 x 10(-8) M. A high analysis rate of 22 samples per hour was obtained. The proposed method has been applied successfully to the determination of scavenging effects of thiourea and vitamin C on hydroxyl radicals as well as to the evaluation of antioxidant capacities of some natural food.     

Phosphite determination in fertilizers after online sequential sample preparation in a flow injection system

A flow injection spectrophotometric system is proposed for phosphite determination in fertilizers by the molybdenum blue method after the processing of each sample two times on-line without and with an oxidizing step. The flow system was designed to add sulfuric acid or permanganate solutions alternately into the system by simply displacing the injector-commutator from one resting position to another, allowing the determination of phosphate and total phosphate, respectively. The concentration of phosphite is obtained then by difference between the two measurents. The influence of flow rates, sample volume, and dimension of flow line connecting the injector-commutator to the main analytical channel was evaluated. The proposed method was applied to phosphite determination in commercial liquid fertilizers. Results obtained with the proposed FIA system were not statistically different from those obtained by titrimetry at the 95% confidence level. In addition, recoveries within 94 and 100% of spiked fertilizers were found. The relative standard deviation (n = 12) related to the phosphite-converted-phosphate peak alone was 相似文献