Development of a Green Chromatographic Method for Simultaneous Determination of Food Colorants

A green chromatographic method for the successful separation and determination of eight synthetic food colorants (Tartrazine E 102, Quinoline Yellow E 104, Sunset Yellow E 110, Carmoisine E 122, Ponceau 4R E 124, Allura Red E 129, Indigo Carmine E 132 and Brilliant Blue E 133) was developed. A C8 stationary phase was used and the mobile phase was a mixture of 50 mM phosphate buffer at pH 7 containing triton X-100 (0.25% v/v). The method was validated as regards its selectivity, linearity, precision, accuracy, limit of detection (LOD) and quantification (LOQ). LOD of colorants varied between 0.17 μg mL⁻¹ in Allura Red and 1.91 μg mL⁻¹ in Quinoline Yellow. In the case of LOQ, it was ranged from 0.52 in the Allura Red to 5.79 in the Quinoline Yellow. The method applicability was verified by the determination of colorants present in 22 samples. The 15 samples were only unicolor and the color concentration in these samples varied from 18.426 ± 0.100 to 610.390 ± 4.711 ppm. The method can be used successfully to the determination of binary and ternary color food and drug samples too. This method provides substantial green benefits without using organic solvents in extraction procedure and in both liquid and paper chromatographic methods.     

Effect of dynamic high-pressure microfluidization at different temperatures on the antigenic response of bovine β-lactoglobulin

The antigenic response of β-lactoglobulin (β-Lg), treated by dynamic high-pressure microfluidization (DHPM) at different temperatures, was determined by an indirect competitive enzyme-linked immunosorbent assay using polyclonal antibodies from rabbit serum. DHPM treatment causes changes in the protein structure and may influence the antigenicity of β-Lg. DHPM treatment of β-Lg at 90 °C showed significant effects with the antigenic response of 5.2 μg mL⁻¹ (untreated), 45 μg mL⁻¹ (40 MPa), 79 μg mL⁻¹ (80 MPa), 132 μg mL⁻¹ (120 MPa), and 158 μg mL⁻¹ (160 MPa). In combination with temperature treatment (70–90 °C), the antigenic response enhanced as the temperature increased at 160 MPa. The β-Lg antigenicities were about 14, 108, and 158 μg mL⁻¹ at 70, 80, and 90 °C, respectively. However, the influence of DHPM pressures on the antigenic response of β-Lg standards was different. DHPM modified β-Lg standards showed a remarkable increase in antigenicity when treated to 80 MPa. Above 80 MPa, the antigenic response decreased.     

Properties of <Emphasis Type="Italic">Aspergillus subolivaceus</Emphasis> free and immobilized dextranase

Aspergillus subolivaceus dextranase is immobilized on several carriers by entrapment and covalent binding with cross-linking. Dextranase immobilized on BSA with a cross-linking agent shows the highest activity and considerable immobilization yield (66.7%). The optimum pH of the immobilized enzyme is shifted to pH 6.0 as compared with the free enzyme (pH 5.5). The optimum temperature of the reaction is resulted at 60 °C for both free and immobilized enzyme. Thermal and pH stability are significantly improved by the immobilization process. The calculated K _m of the immobilized dextranase (14.24 mg mL⁻¹) is higher than that of the free dextranase (11.47 mg mL⁻¹), while V _max of the immobilized enzyme (2.80 U μg protein⁻¹) is lower than that of the free dextranase (11.75 U μg protein⁻¹). The immobilized enzyme was able to retain 76% of the initial catalytic activity after 5.0 cycles.     

Beverage iodine levels in Germany

Sales of iodized table salt (household use, bulk containers) have been flat in Germany since 1996/1997. Nevertheless, the amount of iodide excreted with the urine has been steadily increasing in the German population since 1993. We investigated the beverage iodine levels as possible source of the increased input of iodine into the food chain. No systematic studies on this topic have yet been carried out in Germany. A total of 510 samples were drawn from large grocery discounters who supply all German states with their products (35% fruit and vegetable juices, 35% beers, 10% pooled milk and yoghurt drinks, 10% mineral water, 10% soda). (1) Water-based beverage samples are prepared by solid-phase extraction and iodine concentration is determined by HPLC and electrochemical detection, (2) the samples of milk and milk products are prepared by trichloroacetic acid precipitation and centrifugation, and iodide concentration is determined by HPLC and electrochemical detection. Because the iodine concentrations in the beverage samples showed a left-skewed distribution, all expressed values are median values. Low iodine concentrations were observed in the water-based beverages: fruit and vegetable juices 2.92 μg I⁻/l, beers 2.91 μg I⁻/l, mineral waters 0.5 μg I⁻/l, sodas 0.5 μg I⁻/l. The median value measured in pooled milk and its derived products was 117 μg I⁻/l and 126 μg/l, respectively. As expected, the iodine was lower in summer milk than winter milk (and respectively). Although not statistically significant, beers from Northern Germany had higher concentrations of iodine than from Southern Germany (). In conclusion, based on our results, milk and milk products (117 μg I⁻/l and 126 μg/l, respectively) contribute significantly to the increased nutritional iodine supply of the German population.     

Simultaneous and Direct Determination of As, Bi, Pb, Sb, and Se and Co, Cr, Cu, Fe, and Mn in Milk by Electrothermal Atomic Absorption Spectrometry

Tungsten permanent modifier with coinjection of Pd(NO₃)₂ and W–Ru permanent modifiers are proposed for the direct and simultaneous determination of As, Bi, Pb, Sb, and Se (group 1) and Co, Cr, Cu, Fe, and Mn (group 2), respectively, in milk by graphite furnace atomic absorption spectrometry. The performance of modifiers was evaluated by means of thermal behavior of analytes, sensitivity, atomic signal profile, repeatability, graphite tube lifetime, and background intensity. An air-assisted pyrolysis step was necessary to quantitative elimination of the organic matter. After methods optimization, 14 commercial milk samples were analyzed. The found concentrations of As, Bi, Pb, Sb, Se, Co, and Cr were lower than their limit of detection (2.13, 2.21, 1.49, 1.63, 2.05, 1.0, and 1.2 μg L⁻¹, respectively). Concentrations of Cu, Fe, and Mn were in the 1.58–5.74 μg L⁻¹, 9.79–49.3 μg L⁻¹, and 2.25–4.08 μg L⁻¹ intervals, respectively. The limits of detection for Cu, Fe, and Mn were 1.7, 5.3, and 2.0 μg L⁻¹, respectively. The accuracy of methods was checked after analysis of two milk standard materials. Results for Cr, Cu, Fe, Mn, Pb, and Mn were in agreement with certified values of SRMs at the 95% confidence level. Accuracy was also evaluated by addition–recovery tests and recoveries in the 86–127% range were obtained for all elements. The use of pretreat platform of graphite tubes with W or W–Ru allowed enlarging the lifetime of atomizer in 750 heating cycles.     

Effects of Maillard reaction conditions on the antigenicity of α-lactalbumin and β-lactoglobulin in whey protein conjugated with maltose

The effects of Maillard reaction conditions (weight ratio of protein to sugar, temperature and time) on the antigenicity of α-lactalbumin (α-LA) and β-lactoglobulin (β-LG) in conjugates of whey protein isolate (WPI) with maltose were investigated. Response surface methodology was used to establish models to predict the antigenicity of α-LA and β-LG and find an optimal reaction condition under which the antigenicity of α-LA and β-LG reduces to minimum value. Conjugating WPI with maltose was an effective way to reduce the antigenicity of α-LA and β-LG. The antigenicity of α-LA decreased from 32.25 μg mL⁻¹ to 10.91 μg mL⁻¹. And the antigenicity of β-LG decreased from 272.4 μg mL⁻¹ to 38.17 μg mL⁻¹. Temperature had the greatest effect on the antigenicity of α-LA, while weight ratio of WPI to maltose was the most significant factor on the antigenicity of β-LG.     

Co-production of Lactic Acid and <Emphasis Type="Italic">Lactobacillus rhamnosus</Emphasis> Cells from Whey Permeate with Nutrient Supplements

Lactic acid and cell production from whey permeate by Lactobacillus rhamnosus with different nutrient supplements were investigated in this study. Yeast extract was identified as the most effective nutrient affecting lactic acid production. Increase in inoculum size from 0.05% to 1% (v/v) resulted in a substantial increase in lactic acid productivity from 0.66 to 0.83 g L⁻¹ h⁻¹ (P < 0.001). The optimal temperature for lactic acid production was 37 °C, while the highest cell production was obtained at 42 °C. When whey permeate and yeast extract concentrations were 6.8% (w/v) and 3 g L⁻¹, respectively, lactic acid productivity reached 0.85 g L⁻¹ h⁻¹ after 48-h cultivation, which is 3.40 times of those without nutrient supplements.     

Production of Cell Membrane-Bound α- and β-Glucosidase by Lactobacillus acidophilus

Hydrolytic enzymes, viz. α- and β-glucosidase, were produced from indigenous isolate, Lactobacillus acidophilus, isolated from fermented Eleusine coracana. Production of these enzymes was enhanced by optimizing media using one factor at a time followed by response surface methodology. The optimized media resulted in a 2.5- and 2.1-fold increase in α- and β-glucosidase production compared with their production in basal MRS medium. Localization studies indicated 80% of the total activity to be present in the cell membrane-bound fraction. Lack of sufficient release of these enzymes using various physical, chemical, and enzymatic methods confirmed their unique characteristic of being tightly cell membrane bound. Enzyme characterization revealed that both α- and β-glucosidase exhibited optimum catalytic activity at 50 °C and pH 6.0 and 5.0, respectively. K _m and V _max of α-glucosidase were 4.31 mM and 149 μmol min⁻¹ mL⁻¹ for p-nitrophenyl-α-d-glucopyranoside as substrate and 3.8 mM and 120 μmol min⁻¹ mL⁻¹ for β-glucosidase using p-nitrophenyl-β-d-glucopyranoside as the substrate.     

Micelle-Mediated Extraction and Flame Atomic Absorption Spectrometric Method for Determination of Trace Cobalt Ions in Beverage Samples

A new method has been developed for preconcentration of cobalt at trace levels in beverage samples using calcon carboxylic acid as chelating agent and cetyl pyridinium chloride as an auxiliary ligand and entrapped into Triton X-114 prior to its determination by flame atomic absorption spectrometry (FAAS). The main parameters affecting cloud point extraction (CPE) efficiency such as pH, concentration of the complexing agent, cationic and nonionic surfactant concentration, salt effect, the equilibrium time, and temperature were investigated and optimized. After optimization of the CPE conditions, a preconcentration factor of 60, an enhancement factor of 106, and a detection limit of 0.20 μg L⁻¹ by (R ² = 0.9978) were obtained from a calibration curve constructed in the range of 0.7–100 μg L⁻¹. The proposed preconcentration procedure was successfully applied to the determination of cobalt ions in some real samples including natural drinking water, tap water, and beer and wine samples. The accuracy and validity of the proposed CPE/FAAS method was tested by means of five repeated analysis of reference standard materials (TM-253, a low level fortified water standard for trace elements). A good agreement between analytical results (28.8 and 28.5 μg L⁻¹ with calibration curve and standard addition curve method, respectively) and certified value (27.9 μg L⁻¹) for Co (p < 0.05) were obtained and verified by means of calibration curve and standard addition curve method using CPE procedure.     

Quantitative Determination and Confirmation of Five Synthetic Glucocorticoid Residues in Milk Powder by Gel Permeation Chromatography–Liquid Chromatography–Tandem Mass Spectrometry

A new method for multi-residue determination of five synthetic glucocorticoids (betamethasone, dexamethasone, prednisone, prednisolone, and hydrocortisone) in milk powder is developed. The glucocorticoids in the samples were extracted with tert-butyl methyl ether under ultrasonication incubation, and then cleaned up through gel permeation chromatography. After C18 LC gradient elution separation using acetonitrile with 0.1% formic acid as a mobile phase, the eluents were determined by liquid chromatography–tandem mass spectrometry with multiple reaction monitoring and positive ionization modes. The effective separation for the five glucocorticoids was achieved. The limit of quantification of the method for testing five glucocorticoids in milk powder was in the range from 0.2 to 0.5 μg kg⁻¹, which was lower than the maximum residue limits established by European Union for glucocorticoids in foods. Experiments on spiked samples of milk powder showed that the mean recoveries at addition level of 1.0, 3.0, and 5.0 μg kg⁻¹ were in the range of 71.2% and 103%, and the relative standard deviation ranged from 4.7% to 16.8%. The calibration curves for these drugs between 1.0 and 100 μg l⁻¹ showed good linearity, with correlation coefficient (r) more than 0.999. The real sample test showed that this method is sensitive and accurate. It can be used for qualitative and quantitative determination of studied glucocorticoid residues in milk powder samples.     

A Sensitive, Selective New Analytical Reagent, 2, 6-Diacetylpyridine bis-4-phenyl-3-thiosemicarbazone for Extractive Spectrophotometric Determination of Mo(VI) in Food Samples

Synthesized a new reagent 2, 6-Diacetylpyridine bis-4-phenyl-3-thiosemicarbazone (2, 6-DAPBPTSC) characterized and is proposed as a sensitive and selective analytical reagent for the extractive spectrophotometric determination of molybdenum(VI) at pH 3.5 to form a yellowish orange colored 1:1 chelate complex. The absorbance was measured at a maximum wavelength, 500 nm. This method obeys Beer’s law in the concentration range 0.90–9.00 μg mL⁻¹ and the correlation coefficient of Mo(VI)–2,6-DAPBPTSC complex is 0.954, which indicates an adequate linearity between the two variables with good molar absorptivity and Sandell’s sensitivity, 1.212 × 10⁴ L mol⁻¹cm⁻¹, 0.0079 μg cm⁻², respectively. The precision and accuracy of the method is checked with calculation of relative standard deviation (n = 5), 0.894% and the detection limit value is 0.0056 μg mL⁻¹. The instability constant of the method has been calculated by Asmus’ method as 6.476 × 10^–5, at room temperature. The interfering effect of various cations and anions has also been studied. The method was successfully applied for the determination of Mo(VI) in food and water samples. The validity of the present method was evaluated in terms of Student ‘t’ test and Variance ‘f’ test, which indicates the significance of the present method an inter comparison of the experimental values, using atomic absorption spectrometer.     

Total Mercury, Inorganic Mercury and Methyl Mercury Determination in Red Wine

This study deals with the development of a method for total Hg, inorganic Hg and methylmercury (MeHg) determination in red wine by using flow injection-cold vapour generation–inductively coupled plasma mass spectrometry (FI-CVG-ICP-MS) and gas chromatography-ICP-MS (GC-ICP-MS). For Hg speciation analysis, a derivatization step was carried out using a 1% (m/v) sodium tetraphenylborate (NaBPh₄) solution, followed by extraction of Hg species and their quantification by GC-ICP-MS. The main parameters evaluated were the make-up gas flow rate, volume of the NaBPh₄ solution, time for derivatization reaction/analyte extraction and solvent used for Hg species extraction. Accuracy was evaluated by analyte recovery, whereas recoveries ranged from 99% to 104% for Hg(II) and MeHg. The limits of detection (LODs) for Hg(II) and MeHg were 0.77 and 0.80 μg L⁻¹, respectively. Wine from Argentina, Brazil, Chile and Uruguay were analysed. The wine samples were also acid digested for total Hg determination by FI-CVG-ICP-MS. The LOD of the method used for total Hg determination was 0.01 μg L⁻¹. The concentrations of Hg species in red wine measured by GC-ICP-MS were lower than the respective LODs. Only total Hg was detected in the analysed samples, where the highest concentration of Hg found was 0.55 ± 0.02 μg L⁻¹.     

Determination of Rosmarinic Acid by High-Performance Liquid Chromatography and Its Application to Certain Salvia Species and Rosemary

This study describes a method development for the determination of rosmarinic acid (RA) by using a gradient high-performance liquid chromatography (HPLC) and its application to certain plant materials. The analysis was performed by utilizing a two solvents system [A: methanol/water/formic acid (10:88:2; v:v:v); B: methanol/water/formic acid (90:8:2; v:v:v)] on a reverse-phase column. The flow rate and injection volume were 1 ml min⁻¹ and 10 μl, respectively. Signals were detected at 280 nm. In addition, an internal standard (IS) technique was applied for the analysis of RA to increase precision, and propylparaben was employed for this purpose. The repeatability results as RSD% were 1.66, 1.17 and 1.26 for intra-day and 1.38 was for inter-day with the employment of (3.67 × 10⁻⁵ M) RA. A limit of linearity (LOL) was observed in a wide (1.13 × 10⁻⁵–5.65 × 10⁻⁴ M) concentration range. Linearity parameters were also examined in the range of 5.95 × 10⁻⁶–7.14 × 10⁻⁵ M RA, and very good correlation was observed. The limit of detection (LOD) and limit of quantification (LOQ) (for inter-day) were 1.60 × 10⁻⁶ M (signal/noise [S/N] = 3.3) and 4.80 × 10⁻⁶ M (S/N = 10), respectively. The method was applied to the extracts of certain Lamiaceae plants (Salvia candidissima Vahl. subsp. candidissima, S. sclarea L., S. verticillata L. subsp. verticillata and R. officinalis L.), and reasonable results were obtained.     

Development of an Analytical Method Based in the Slurry Sampling for Iron Determination in Fortified Milk Powder by HR-CS FAAS

The use of slurry sampling as the procedure for sample preparation provides simplicity, speed, and low consumption of reagents in analytical methods. In this paper, a method based on slurry sampling for the determination of iron in samples of fortified milk powder by high-resolution continuum source flame atomic absorption spectrometry was developed. Multivariate design techniques were applied for the optimization of experimental conditions of the method for a sample mass of 100 mg, final volume of slurry of 10 mL, and using the absorbance signal as response. Initially, a two-level full factorial design was used for the preliminary evaluation of the variables involved in slurry preparation: concentration hydrochloric acid, Triton X-100 concentration, and sonication time. Then, the Doehlert matrix was applied for the determination of the critical conditions: 2.5 mol L⁻¹ hydrochloric acid and sonication time of 20 min. External calibration technique with aqueous standard was used for the quantification of iron. This way, the method allowed iron determination with limits of detection and quantification of 0.9 and 3.0 μg g⁻¹, respectively. The precision expressed as the relative standard deviation was evaluated under repeatability and reproducibility conditions, being 3.2% and 4.0%, respectively. Addition/recovery test was used for assessing the accuracy of the method, and the recovery values achieved were in the range of 90–110%. The method was applied for iron determination in eight samples of fortified milk powder, and the obtained concentrations varied from 95.4 to 295.6 μg g⁻¹. The results were compared with those obtained after acid digestion, and no significant difference was observed applying t test at the 95% confidence level.     

Ultrasonically Assisted Extraction of Rutin from <Emphasis Type="Italic">Artemisia selengensis</Emphasis> Turcz: Comparison with Conventional Extraction Techniques

Ultrasonically assisted extraction (UAE) followed by high performance liquid chromatography (HPLC) analysis method for the fast extraction and determination of rutin in Artemisia selengensis Turcz has been developed. Artemisia selengensis Turcz has been used as food and herbal medicine for thousands of years in China. Rutin is one of the main active ingredients of this plant. The extraction of rutin from Artemisia selengensis Turcz was investigated by UAE. Special emphasis has been given to optimize the extraction conditions which were those with 90:10 (v/v) methanol–ethanol as solvent, 30:1 liquid–solid ratio, and 40 min extraction time. In order to show the superiority of UAE, other extractions were investigated, including microwave-assisted extraction, reflux extraction, and marinated extraction. The results showed that UAE was most suitable for the extraction of rutin in Artemisia selengensis Turcz because of its high extraction efficiency. Reversed phase-HPLC with ultraviolet detection was employed for the analysis of rutin in Artemisia selengensis Turcz. Under the optimum conditions, the calibration curve for the analyte was linear in the range of 0.34–20.7 μg mL⁻¹. The mean recovery of rutin was 100.77%, and its relative standard deviation was 0.37% (n = 5). Three kinds of Artemisia selengensis Turcz from different habitats were investigated. The total content of rutin was 9.90, 6.23, 5.56 mg g⁻¹, respectively.     

Determination and Evaluation of the Mineral Composition of Chinese Cabbage (Beta vulgaris)

Inductively coupled plasma optical emission spectrometry was used to determine the elements present in Chinese cabbage (Beta vulgaris). The accuracy of the method was confirmed by analysis of a certified reference material of spinach leaves. The study involved 57 samples that were collected in 13 Brazilian cities. Average concentrations of elements found per gram of Chinese cabbage were as follows: 3.44 mg g⁻¹ sodium, 5.09 mg g⁻¹ potassium, 1.25 mg g⁻¹ phosphorous, 0.85 mg g⁻¹ calcium, 0.49 mg g⁻¹ magnesium, 2.79 μg g⁻¹ manganese, 9.50 μg g⁻¹ iron, 0.74 μg g⁻¹ copper, 14.28 μg g⁻¹ zinc, and 6.44 μg g⁻¹ strontium. Principal component analysis and hierarchical cluster analysis demonstrated that there is no systematic difference in the mineral composition between the cabbage samples that were analyzed.     

Polycyclic aromatic hydrocarbons (PAHs) in traditional and industrial smoked beef and pork ham from Serbia

Smoked beef and pork ham samples were analysed during process of smoking (after packing and storing) for the presence of the 16 EU priority PAHs via Fast GC/HRMS method. This study showed that there are differences in PAH contents between final smoked beef ham samples from traditional smokehouse (TS) (3.9 μg kg⁻¹) and industrial smokehouse (IS), (1.9 μg kg⁻¹). Also there is a difference in PAH contents in final smoked pork ham samples (4.9 μg kg⁻¹, TS; 4.2 μg kg⁻¹, IS). In beef and pork ham samples from the same smokehouse different PAH contents were observed during smoking. The highest content of examined PAHs in all beef and pork ham samples during smoking showed benzo[c]fluorene (BcL) (beef ham: from 0.3 μg kg⁻¹ to 1.5 μg kg⁻¹; pork ham: from 0.2 μg kg⁻¹ to 2.1 μg kg⁻¹).The maximum level for benzo[a]pyrene (BaP) of 5 μg kg⁻¹ in smoked meat products was not exceeded in any samples. Correlation statistic analysis (P < 0.05) of obtained contents from samples both from TS and IS showed that BaP is a good marker both for 16 EU priority PAHs and 12 IARC probably and possibly carcinogenic PAHs (IS: R _BaP/Σ16PAHs = 0.95, R _BaP/Σ12PAHs = 0.96; TS: R _BaP/Σ16PAHs = 0.71, R _BaP/Σ12PAHs = 0.88).     

An approach to improve ACE-inhibitory activity of casein hydrolysates with plastein reaction catalyzed by Alcalase

The preparation method of casein hydrolysates with high ACE-inhibitory activity was studied by Alcalase-catalyzed hydrolysis coupled with plastein reaction. Casein hydrolysates with an IC₅₀ value of about 47 μg mL⁻¹ were first prepared by hydrolysis of casein with Alcalase and then modified with plastein reaction catalyzed by the same enzyme. The impacts of four reaction conditions on plastein reaction of casein hydrolysates were studied, and then optimal conditions were determined using response surface methodology with the decrease of free amino groups in the reaction mixture as response. When the concentration of casein hydrolysates was fixed at 35% by weight, the maximum decrease of free amino groups in the reaction mixture of 181.8 μmol g⁻¹ proteins was obtained. The optimum conditions for the above decrease were found to be an E/S ratio of 7.7 kU g⁻¹ proteins, reaction temperature of 42.7 °C and reaction time of 6 h. Analysis results showed that ACE-inhibitory activity of casein hydrolysates prepared could be improved significantly by plastein reaction. When casein hydrolysates were modified by plastein reaction, with a decrease of free amino groups in the mixture of about 154.7 μmol g⁻¹ proteins and 181.8 μmol g⁻¹ proteins, their IC₅₀ values could be decreased to 0.6 and 0.5 μg mL⁻¹.     

Sensitive Determination of Decoquinate in Milk by High-Performance Liquid Chromatographic Coupled to Laser-Induced Fluorescence Detection

A new and sensitive method based on high performance liquid chromatography with laser-induced fluorescence detection has been developed for the determination of decoquinate in milk. Laser source was obtained with a He–Cd laser using a continuous excitation wavelength at 325 nm. Decoquinate exhibits moderate fluorescence, but it is increased using Ca(NO₃)₂ in the mobile phase. The chromatographic separation was performed on a Luna C₁₈ 5-mm reversed phase column, which solves the broadening of peaks and peak tailing compared with other columns tested. The mobile phase, delivered at 1 ml min⁻¹, consisted of methanol–calcium nitrate (0.025 M)–acetonitrile (83/13/4 v/v/v). Decoquinate was successfully cleaned up from milk by solid-phase extraction using C₁₈ cartridges. The method was found to be linear between 0.16 and 16.33 ng ml⁻¹. The results of recovery studies were found to be satisfactory; an average recovery rate of 88.7% was obtained. The LOQ of decoquinate in milk was 0.16 ng ml⁻¹. The intraday relative standard deviation (RSD) was 4%, and interday assay gave an RSD of 4.4%.     

Chemiluminescence Detection of 2,4,5-Trichlorophenoxy Acetic Acid in Apple Juice by Digital Image Analysis

An image-based detection of an enhanced chemiluminescence enzyme-linked immunosorbent assay was developed for 2,4,5-trichlorophenoxy acetic acid with an anti-rabbit secondary antibody conjugated to peroxidase (goat anti-rabbit IgG–HRP). Data acquisition on microtiter wells is performed using a low-cost charge-coupled device camera for capturing images. The standard curve was produced for 0.01–5,000 ng mL⁻¹ of 2,4,5-trichlorophenoxy acetic acid. The minimum detectable concentration was 24 pg mL⁻¹, and the relative standard deviation was 5.5% (n = 10) for a 5 ng mL⁻¹ sample concentration. Similar sensitivity was obtained with enzyme-linked immunosorbent assay using the same polyclonal antibody and data acquisition by a spectrofluorometer. This method was applied to apple juice with recoveries between 95% and 111% and relative standard deviation between 6.1% and 11.3%.