Determination of sulfite in dried garlic by reversed-phase ion-pairing liquid chromatography with post-column detection

A method is described for determining sulfite in dried garlic. Garlic is extracted with an HCl solution to inhibit the formation of allicin, which interferes with the determination of sulfite. After cleanup of the extract on a C18 solid-phase extraction column, sulfite is converted to hydroxymethylsulfonate (HMS) by adding formaldehyde and heating to 50 degrees C. HMS is determined by reversed-phase ion-pairing liquid chromatography with post-column detection. The post-column reaction system consists of the addition of KOH to convert HMS to sulfite ion, followed by the addition of 5,5'-dithiobis(2-nitrobenzoic acid) to produce 5-mercapto-2-nitrobenzoic acid which is detected spectrophotometrically at 450 nm. Background levels in unsulfited dried garlic equivalent to < 20 ppm SO2 were found. Recoveries of HMS from spiked garlic averaged 94.8% with a coefficient of variation of 3.8%. Sulfite was found in 13 of 21 samples of dried garlic produced in China, with sulfite ranging from 114 to 445 ppm. Sulfite was found in 60% of commercial dried garlic products purchased locally. The suitability of the Monier-Williams method for determining sulfite in garlic is discussed.     

High-performance ion-pair chromatography method for simultaneous analysis of alliin,deoxyalliin, allicin and dipeptide precursors in garlic products using multiple mass spectrometry and UV detection

The quality of garlic and garlic products is usually related to their alliin content and allicin release potential. Until now no analytical method was able to quantify simultaneously allicin, its direct precursor alliin (S-allyl-L-cysteine sulfoxide), SAC (S-allyl-L-cysteine) as well as various dipeptides that apparently serve as storage compounds in garlic. It is well known that all these intermediates are involved in the allicin biosynthetic pathway. A simple and rapid HPLC method suitable for routine analysis was developed using eluents containing an ion-pairing reagent. Particularly, heptanesulfonate as ion-pairing reagent guarantees a sufficient separation between alliin and the more retained dipeptides at very low pH. Allicin was eluted after 18 min on a 150 x 3 mm column. Synthetic reference compounds were characterized by the same chromatographic method using a diode-array UV detector and an ion trap mass spectrometer (electrospray ionization) in the multiple MS mode. In routine analysis of garlic bulbs, powders and other products, the diode-array detector is sufficient for a relevant quantification. Our method has been used in studies to improve the quality of garlic and its derived products.     

天然大蒜油及合成大蒜素的气相色谱-质谱分析

利用气相色谱-质谱联用方法对天然大蒜油和合成大蒜素进行了分析和鉴定，给出了其主要组分及分布情况。对两者质谱图进行了比较，发现在合成大蒜素中含有微量的氯丙烯，据此发展了一种快速灵敏的分析方法，用来判断实际大蒜油样品中是否含有合成大蒜素。     

大蒜素的检测及大蒜辣素分解机理的探讨

通过正交试验方法,研究了硫酸钡吸光比浊法测定大蒜中大蒜素含量的最佳实验条件。在选定实验条件下,体系的吸光度与硫酸根离子浓度呈线性关系,相关系数r为0.9985,相对标准偏差(RSD)为0.86%。采用高效液相色谱法对样品中二烯丙基二硫醚进行定量检测,其检测结果与吸光比浊法的结果具有一定的互补对照性。在此基础上采用红外光谱法、气相色谱-质谱法对大蒜素中含硫活性成分进行定性分析,并将四种方法结果进行对比,对大蒜辣素过热分解的机理进行了探讨,为大蒜素中不同含硫化合物之间的转化提供了依据。     

Validated liquid chromatographic method for quantitative determination of allicin in garlic powder and tablets

In the present study, an RP high performance liquid chromatographic method was developed and validated for the determination of allicin in garlic powder and tablets. Chromatographic separation was carried out on an RP-18(e )column (125 mm x 4 mm), using a mobile phase, consisting of methanol-water (50:50 v/v), at a flow rate of 0.5 mL/min and UV detection at 220 nm. Ethylparaben was used as the internal standard. The assay was linear for allicin concentrations of 5.0-60.0 microg/mL. The RSD for precision was <6.14%. The accuracy was above 89.11%. The detection and quantification limits were 0.27 and 0.81 microg/mL, respectively. This method was used to quantify allicin in garlic powder samples. The results showed that the method described here is useful for the determination of allicin in garlic powder and tablets.     

Determination of sulfite in beer samples using an amperometric fill and flow channel biosensor employing sulfite oxidase

A simple method is described to determine sulfite in beer samples using a fill and flow channel biosensor. A droplet of sample is placed into the inlet of a rectangular flow cell and begins to flow through the channel by capillarity. The flow is maintained and controlled by a porous outlet plug of defined porosity. In a rectangular flow cell, the sample solution flows through three consecutive zones: over a predictor electrode, an enzyme layer and a detector electrode. Together these three zones enable the differentiation between current due to sulfite and current due to other electroactive species in the sample. The predictor electrode is located upstream, and on the opposite channel wall to the enzyme layer and detector electrode, and is poised at the same potential (+0.65 V versus Ag/AgCl) as the detector electrode. On this electrode, the current contribution from all species in the sample solution that are oxidized at that potential is determined. The enzyme layer contains sulfite oxidase, which, in the process of oxidizing sulfite, produces hydrogen peroxide, which itself is reduced by excess sulfite. The current at the downstream detector electrode is therefore different from that at the predictor electrode as a result of the enzyme reaction and the difference of the currents, corrected for the dimensions of the electrodes, is proportional to the concentration of sulfite. The method enables a straightforward correction of the interfering current at the detector electrode and a determination of the analyte concentration. The effect of interferences from ascorbic acid, ethanol, sorbic acid and tartaric acid in the detection of sulfite is efficiently removed. The concentration of sulfite in a sample of beer measured by the biosensor is equivalent to that measured using a reference method based on the AOAC-recommended Monier-Williams method.     

Simultaneous kinetic determination of sulfite and sulfide using artificial neural networks

Artificial neural networks (ANNs) are proposed for the determination of sulfite and sulfide simultaneously. The method is based on the reaction between Brilliant Green (BG) as a colored reagent and sulfite and/or sulfide in buffered solution (pH 7.0) and monitoring the changes of absorbance at maximum wavelength of 628 nm. Experimental conditions such as pH, reagents concentrations, and temperature were optimized and training the network was performed using principal components (PCs) of the original data. The network architecture (number of input, hidden and output nodes), and some parameters such as learning rate (η) and momentum (α) were also optimized for getting satisfactory results with minimum errors. The measuring range was 0.05-3.6 μg ml⁻¹ for both analytes. The proposed method has been successfully applied to the quantification of the sulfite and sulfide in different water samples.     

Electrocatalytic Oxidation and Determination of Sulfite with a Novel Copper‐Cobalt Hexacyanoferrate Modified Carbon Paste Electrode

The electrocatalytic oxidation of sulfite has been studied at a stable electroactive thin film of copper‐cobalt hexacyanoferrate (CuCoHCF) hybrid electrodeposited on a carbon paste electrode (ECMCPE). A linear range of 5 μM to 5 mM of sulfite, with an experimental detection limit of 1 μM, was obtained using the cyclic voltammetric method. The oxidation of sulfite showed no significant fouling effect on the modified electrode surface at sulfite concentrations below 5 mM. The proposed modified electrode exhibited several attractive features, including simple preparation, fast response, good stability and repeatability, and could be applied to sulfite determination in real samples.     

Sulfite oxidase biosensors based on tetrathiafulvalene modified screen-printed carbon electrodes for sulfite determination in wine

Screen-printed carbon electrodes have been modified with tetrathiafulvalene and sulfite oxidase enzyme for the sensitive and selective detection of sulfite. Amperometric experimental conditions were optimized taking into account the importance of quantifying sulfite in wine samples and the inherent complexity of these samples, particularly red wine. The biosensor responds to sulfite giving a cathodic current (at +200 mV vs screen-printed Ag/AgCl electrode and pH 6) in a wide concentration range, with a capability of detection of 6 μM (α = β = 0.05) at 60 °C. The method has been applied to the determination of sulfite in white and red samples, with averages recoveries of 101.5% to 101.8%, respectively.     

Thermal behaviour and S-O bonding state of alkaline-earth metal sulfites

The thermal behaviours of magnesium sulfite, strontium sulfite and barium sulfite were investigated in the atmospheres of argon and air. the thermal behaviours of magnesium sulfite were different from those of the other two sulfites. The oxidation of magnesium sulfite in air does not occur.The bonding state of the SO ₃ ^2– in each sulfite was compared. The SO ₃ ^2– in magnesium sulfite was coordinated through the sulfur, while those in the other sulfites were coordinated through the oxygen.It appears that the difference in thermal behaviour between magnesium sulfite and the other sulfites depend upon the difference in bonding state of the SO ₃ ^2– We wish to thank Mr. K. Takahashi of MAC Science Inc. for TG and DTA measurements, mass spectra in Ar and his valuable discussions.     

Quantitative Determination of Allicin in Allium sativum L. Bulbs by UPLC

An UPLC method for determination of allicin in garlic bulb powder has been developed and validated. Chromatographic separation and determination were performed on an Acquity UPLC BEH C₁₈ column using a mobile phase of methanol/water (50:50, v/v). Linearity between the allicin concentration and the peak area was good within the concentration ranges 2.04–510 mg L⁻¹. The method was validated over this range for both accuracy and precision. The method was successfully used for the quantitative analysis of allicin in ten garlic varieties.

     

Development and Application of an Immobilized Enzyme Electrode for the Determination of Sulfite in Foods and Feeds

Abstract

An enzyme electrode was developed using the enzyme sulfite oxidase (EC 1.8.3.1) immobilized onto pig intestine. All experimental parameters such as pH, temperature, buffer constituent concentration, etc., were thoroughly investigated and optimized when appropriate. Hydrogen peroxide was monitored amperometrically. The response to sulfite concentration was linear in the range of 5.2 × 10^?6 to 1.0×10^?3M. The proposed method was found to have a correlation coefficient of 0.952 when evaluated versus the standard titration method.     

Electron-bombardment and pyrolysis of meso- and dl-hydrobenzoin cyclic sulfites

Only a minor yield of sulfur dioxide positive ions is eliminated from the meso- and dl-hydrobenzoin cyclic sulfite molecules under electron bombardment. Among the principal fragmentation products for both sulfites are very similar yields of ions with m/e 154 and 126, which contain sulfur most probably attached to a phenyl ring. Positive ions apparently corresponding to those of diphenylacetaldehyde, desoxybenzoin, and stilbene are minor products of the positive-ion fragmentation process of the sulfites. These three compounds along with sulfur dioxide are the products of the pyrolysis of the sulfites at 240-250°. The principal product from the meso-sulfite is desoxybenzoin, and from the dl-sulfite it is diphenylacetaldehyde. Mechanistic details are discussed.     

Allicin,the Odor of Freshly Crushed Garlic: A Review of Recent Progress in Understanding Allicin’s Effects on Cells

The volatile organic sulfur compound allicin (diallyl thiosulfinate) is produced as a defense substance when garlic (Allium sativum) tissues are damaged, for example by the activities of pathogens or pests. Allicin gives crushed garlic its characteristic odor, is membrane permeable and readily taken up by exposed cells. It is a reactive thiol-trapping sulfur compound that S-thioallylates accessible cysteine residues in proteins and low molecular weight thiols including the cellular redox buffer glutathione (GSH) in eukaryotes and Gram-negative bacteria, as well as bacillithiol (BSH) in Gram-positive firmicutes. Allicin shows dose-dependent antimicrobial activity. At higher doses in eukaryotes allicin can induce apoptosis or necrosis, whereas lower, biocompatible amounts can modulate the activity of redox-sensitive proteins and affect cellular signaling. This review summarizes our current knowledge of how bacterial and eukaryotic cells are specifically affected by, and respond to, allicin.     

Electrocatalytic oxidation of sulfite by acetylferrocene at glassy carbon electrode

The electrochemical oxidation of sulfite catalyzed by acetylferrocene (AFc) at a glassy carbon electrode (GCE) in 0.2 M NaClO₄ aqueous solution has been studied by cyclic voltammetry. Although sulfite itself showed a sluggish electrochemical response at the GCE, the response could be enhanced greatly by using AFc as a mediator, which enables a sensitive determination of the substrate (sulfite). The reaction rate constant for catalytic oxidation was evaluated as (7.02 ± 0.05) × 10⁴ M ^?1 s^?1 by chronoamperometry. Experimental conditions that maximize the current efficiency of the electrocatalytic oxidation, such as the pH and both the catalyst (AFc) and substrate (sulfite) concentrations, were also investigated. The electrochemical kinetics of electrocatalytic oxidation of sulfite by AFc has been studied by cyclic voltammetry. In the presence of 5 × 10^?4 M AFc, the oxidation current is proportional to the sulfite concentration and the calibration plot was linear over the concentration range 2 × 10^?4–2.4 × 10^?3 M . This result can be applied in the determination of real samples. Copyright © 2005 John Wiley & Sons, Ltd.     

Determination of sulfite in Oriental herbal medicines

Sulfite was detected in 7 varieties of Oriental herbal medicines (Pueraria radix, Zingiberis rhizoma, Platycodon radix, Adenophora radix, Pinellia tuber, Astragalus radix, and Paeonia radix) on the Korean market. Sulfiting of commercial Oriental herbal medicines by fumigation with burning bituminous coal was simulated, and the accumulation of sulfite was investigated by using fresh Platycodon radix roots obtained from a growing field. The sulfite level reached a plateau in 9 h, and the maximum sulfite level found by the Monier-Williams (MW) method (AOAC 990.28) was 1020 ppm. The sulfite content in the simulated Platycodon radix sample determined by alkali extraction followed by ion-exclusion chromatography with electrochemical detection (AOAC 990.31) was approximately 17% lower on average than the MW results. Free-sulfite levels determined by acid extraction and ion-exclusion chromatography with electrochemical detection were between 19 and 49% of the MW results. The advantages of different methods for sulfite determination and the significance of the results are discussed.     

Determination of total sulfite in wine. Zone electrophoresis-isotachophoresis quantitation of sulfate on a chip after an in-sample oxidation of total sulfite

This work deals with the determination of total sulfite in wine. The determination combines an in-sample hydrogen peroxide oxidation of total sulfite in alkalized wine to sulfate with the separation and quantitation of the latter anion by zone electrophoresis (ZE) on-line coupled with isotachophoresis (ITP) on a column-coupling chip. Sample clean up, integrated into the ITP-ZE separation, eliminated wine matrix in an extent comparable to that provided by a highly selective distillation isolation of sulfite. At the same time, conductivity detection, employed to the detection of sulfate in the ZE stage of the ITP-ZE combination, provided for sulfate the concentration limit of detection corresponding to a 90 microg/l concentration of sulfite in the loaded sample (0.9 microl). Such a detectability allowed a reproducible quantitation of total sulfite when its concentration in wine was 15 mg/l. Formaldehyde binding of free sulfite in wine, included into the pre-column sample preparation, prevented an uncontrolled oxidation of this sulfite form. This step contributed to an unbiased determination of sulfate present in the original wine sample (this determination corrected for the concentration of sulfate determined in the sample after the peroxide oxidation of sulfite to the value equivalent to the total sulfite). The 99-101% recoveries of sulfite, determined for appropriately spiked wine samples, indicate a very good accuracy of the present method. Such a statement also supports excellent agreements of the results of quantitation based on the in-sample peroxide oxidation of the total sulfite (bound sulfite released at a high pH) with those in which this analyte was isolated from wine by distillation (bound sulfite released at a very low pH).     

Allicin induces apoptosis in EL-4 cells in vitro by activation of expression of caspase-3 and -12 and up-regulation of the ratio of Bax/Bcl-2

Garlic (Allium sativum L.; Liliaceae) has been widely demonstrated in the role of cancer prevention, but the specific compound in garlic corresponding to this effect and its mechanisms are not clearly known. Allicin is one of the organic sulphur compounds derived from garlic. In the present study we investigated the anti-proliferative and pro-apoptotic activities of allicin in murine T-lymphocytes (EL-4) and the mechanism of inducing apoptosis in?vitro. The results showed that allicin was effective in inhibiting the proliferation of EL-4 cells in?vitro in a concentration-dependent manner. Further, allicin could induce the formation of apoptotic bodies, nuclear condensation, DNA spallation, and even activated the expression of caspase-3, -12 and cytochrome C (cyt C). Finally, allicin up-regulated the ratio of Bax/Bcl-2 and induced a mitochondrion membrane potential (MMP) decrease. Allicin induced apoptosis in EL-4 cells in a time- and concentration-dependent manner, in which the mitochondrial pathway might play a central role.     

Composition and Stability of a Polynuclear Copper Sulfite Complex

Conditions for the formation of a polynuclear copper sulfite complex were studied by spectrophotometry, conductometry, and chemical analysis. The composition of the forming sulfites was determined, and the equilibrium constant of formation of the polynuclear copper sulfite complex and its dissociation constant were calculated.     

Determination of free and reversibly-bound sulfite in selected foods by high-performance liquid chromatography with fluorometric detection

A rapid and accurate method for measuring low part-per-million levels of free and reversibly-bound sulfites in selected foods by using high-performance liquid chromatography (HPLC) with fluorometric detection was developed. Sulfites were extracted with sodium tetrachloromercurate solution and determined by HPLC-fluorescence spectrometry. During the HPLC analysis, the sample extract was reacted with sodium hydroxide to liberate the reversibly-bound sulfite and subsequently separated from other interferences by a size exclusion column. The effluent was then reacted with o-phthalaldehydelammonium chloride reagent to form a fluorescing 1-sulfonatoisoindole derivative and analyzed by a fluorescence detector. The method has been applied to a variety of foods and food products, with no significant interference encountered in matrixes such as soy products, cabbage, broccoli, brassica, ginger, fungus, mushroom, mandarin peel, potato chips, and biscuits. It was shown to have a broad linear range of 0.01 to 0.4 mg/L as SO2. The spike recoveries of sodium sulfite, sodium metabisulfite, and formaldehyde-sodium bisulfite adduct at the 5 to 10 mg/kg level in the tested food matrix were within 80-120%, and the limit of detection was 5 mg/kg. Laboratory of Government Chemist Reference Material LGC7111 (potato powder) was used to assess the accuracy of the method. The expanded measurement uncertainty of the method at 95% confidence level was estimated to be 17%.