Carbohydrate metabolism dynamic in chlorpropham- and 1,4-dimethylnaphthalene-treated potatoes and its effect on the browning of French fries.

The use of a sprout suppressor is crucial for the use of potatoes beyond their natural dormancy period. The main sprout inhibitor used on a commercial scale, chlorpropham (CIPC), is becoming increasingly limited owing to its toxicity. Therefore, we evaluated the effectiveness of 1,4-dimethylnaphthalene (1,4-DMN) compared to CIPC in controlling sprouting and maintaining the quality of potato, Solanum tuberosum 'Asterix', during cold storage. Treatment with 1,4-DMN reduced fresh weight loss and controlled the number and length of sprouts comparable to CIPC. Compared to the control, both sprouting inhibitors led to higher starch and lower reducing sugar contents, and the tubers retained the recommended quality for industrial processing. After frying, less browning was observed in French fries obtained from 1,4-DMN- or CIPC-treated tubers. We ascertain that 1,4-DMN besides being an efficient sprouting inhibitor and alternative to CIPC, it contributes to maintaining the quality of French fries after cold storage.

Fate of Chlorpropham during High-Temperature Processing of Potatoes.

Chlorpropham is a widely used sprouting inhibitor applied on potatoes during their storage. Currently, severe concerns are raised regarding the potential formation of 3-chloroaniline from chlorpropham during heat treatment. The reactions degrading the molecule in the matrix are quite complex under harsh processing conditions, and a molecular investigation is thus challenging. This study aims to decipher the reaction pathways and to discover new metabolites in typical high-temperature food-processing steps. For this purpose, potatoes were treated with 14C-radiolabeled chlorpropham, stored for up to 6 months, and subjected to the traditional preparation steps of boiling, frying, and baking. A quantification method including an acidic hydrolysis was developed for analysis of free and bound analytes. All conducted processing steps led to a substantial mitigation of chlorpropham residues in the consumable products. Of the residues, 17 ± 6% remained in boiled tubers, while 27 ± 3 and 22 ± 3% remained in the fried and baked products, respectively. Chlorpropham was transferred into the surrounding media (boiling water, frying oil, and air, respectively). 3-Chloroaniline was only (raw tubers) or predominantly (processed tubers) present as a bound analyte and was shown to form during storage but not during processing. Additionally, nonextractable and nonquantified residues were detected in the baked and in the long-term-stored tubers after processing. Future studies will have to balance beneficial (mitigating) and potentially hazardous aspects of these results. By transferring the 14C-food-processing approach to a variety of substances, ingredients, and processes, it will be possible to further understand chemical reactions in food processing, finally leading to safer food.

Biodegradation of chlorpropham and its major products by Bacillus licheniformis NKC-1.

Chlorpropham [isopropyl N-(3-chlorophenyl) carbamate] (CIPC), an important phenyl carbamate herbicide, has been used as a plant growth regulator and potato sprout suppressant (Solanum tuberosum L) during long-term storage. A bacterium capable of utilizing the residual herbicide CIPC as a sole source of carbon and energy was isolated from herbicide-contaminated soil samples employing selective enrichment method. The isolated bacterial strain was identified as Bacillus licheniformis NKC-1 on the basis of its morphological, cultural, biochemical characteristics and also by phylogenetic analysis based on 16S rRNA gene sequences. The organism degraded CIPC through its initial hydrolysis by CIPC hydrolase enzyme to yield 3-chloroaniline (3-CA) as a major metabolic product. An inducible 3-CA dioxygenase not only catalyzes the incorporation of molecular oxygen but also removes the amino group by the deamination yielding a monochlorinated catechol. Further, degradation of 4-chlorocatechol proceeded via ortho- ring cleavage through the maleylacetate process. 3-Chloroaniline and 4-chlorocatechol are the intermediates in the CIPC degradation which suggested that dechlorination had occurred after the aromatic ring cleavage. The presence of these metabolites has been confirmed by using ultra-violet (UV), high-performance liquid chromatography (HPLC), thin layer chromatography (TLC), Fourier transmission-infrared (FT-IR), proton nuclear magnetic resonance (1H NMR) and gas chromatography-mass (GC-MS) spectral analysis. Enzyme activities of CIPC hydrolase, 3-CA dioxygenase and chlorocatechol 1, 2-dioxygenase were detected in the cell-free-extract of the CIPC culture and are induced by cells of NKC-1 strain. These results demonstrate the biodegradation pathways of herbicide CIPC and promote the potential use of NKC-1 strain to bioremediate CIPC-contaminated environment with subsequent release of ammonia, chloride ions and carbon dioxide.

Determination of chlorpropham (CIPC) residues, in the concrete flooring of potato stores, using quantitative (HPLC UV/VIS) and qualitative (GCMS) methods.

Isopropyl-N-(3-chlorophenyl) carbamate (CIPC, common name Chlorpropham) is commonly used for post-harvest sprout inhibition in stored potatoes. It is applied as a thermal fog which results in loss to the fabric of the store and the atmosphere. Recently, there have been concerns in the United Kingdom because of cross contamination of other crop commodities that were stored in buildings with a history of CIPC usage. This cross contamination may have occurred because of retained residues in the fabric of the stores. The retention of CIPC in concrete is poorly understood; therefore the requirement for a robust analytical method for the detection and quantification of CIPC in concrete is a critical first step in tackling this problem. A method using High-Performance Liquid Chromatography with ultraviolet detection (HPLC UV/VIS) was validated. CIPC recoveries at three concentration levels (0.4, 4.0 and 40.0 µg g-1) were in the range of 90.7-97.0% with relative standard deviations between 2.14 and 3.01%. The limits of detection and quantification were 0.03 and 0.1 µg g-1, respectively. This study confirmed that CIPC was persistent in concrete to a depth of 4 cm, with >90% within the top 1 cm of the flooring.

Complexation of chlorpropham with hydroxypropyl-ß-cyclodextrin and its application in potato sprout inhibition.

The effect of hydroxypropyl-ß-cyclodextrin (HPßCD) on the improvement of chlorpropham (CIPC) as a potato sprout inhibitor was investigated. The formation of complex was confirmed by FT-IR spectra, thermoanalysis, (1)H NMR and ROESY. The stoichiometry and stability constant were determined by Job's plot and phase solubility studies, respectively. The inclusion complex CIPC·HPßCD has exhibited different properties from CIPC. The obtained inclusion complex was found to significantly improve the water solubility, thermal stability and dissolution rate of CIPC. In addition, the complex displayed a better effect on sprout inhibition.

Direct detection of chlorpropham on potato skin using desorption electrospray ionization.

Most pesticides, herbicides and other plant treatment agents are applied to the crop surface. Direct mass spectrometric methods, such as desorption electrospray ionization (DESI), offer new ways to analyze plant samples directly and rapidly. A strategy for the development and optimization of a DESI method for the direct determination of chemicals on complex surfaces is described. Chlorpropham (CP) was applied to potato surfaces as an example for a crop protection agent and analyzed using a self-made DESI source. Aspects such as instrument selectivity, sensitivity and reproducibility were investigated. The MS(4) fragmentation pattern of CP was analyzed to achieve the necessary detection selectivity, and is discussed in detail. Similar fragmentation was found in the ESI and DESI mass spectra, indicating that the mechanisms of ESI and DESI are closely related. A DESI method for semi-quantification of CP on potatoes was developed. Detection limits of 6.5 µg/kg were found using MS/MS. The reproducibility, in the range of 12% (signal variation), appears to be sufficient for semi-quantitative measurements.

Micronucleation by mitosis inhibitors in developing microspores of Spathiphyllum wallisii Regel.

KEY MESSAGE : We developed an efficient protocol for chromosome scattering in Spathiphyllum microspores. The effects of plant material, developmental age, genotype and antimicrotubular toxin type, exposure and concentration were evaluated. Asymmetric hybridization through microprotoplast-mediated chromosome transfer (MMCT) is a known method for overcoming sexual breeding barriers between distantly related plant species. To obtain microprotoplasts, it is necessary to induce mass micronucleation either in somatic or gametic cells. We have tested the efficiency for micronuclei induction of five mitosis inhibitors, amiprophos-methyl (APM), butamiphos (BUT), chlorpropham (CIPC), oryzalin (ORY) and propyzamide (PRO), on developing microspores of diploid Spathiphyllum wallisii Regel. Besides the used toxins, also the effect of their concentrations and incubation period as well as plant genotypes and material was tested. We observed micronuclei (MNi) in pollen mother cells, dyads and tetrads as well as other abnormalities such as ball metaphases and chromosome bridges. The flower position on the spadix and the type of starting material (dissected anthers vs. complete spadices) did not significantly influence micronucleation frequencies. The highest micronucleation index of 86 % was obtained in microspores treated with 10 µM ORY during 72 h. All six genotypes tested formed micronuclei after this particular treatment, although the efficiency varied between cultivars. Next to ORY, CIPC was also a very efficient MNi inducer. The average number of MNi found in micronucleated cells varied between 1.67-6.44 for CIPC and 0.83-5.50 for ORY. The maximal number of MNi observed was 12 for CIPC and 9 for ORY. Our results demonstrate that CIPC and ORY can be applied for mass micronucleation on developing microspores of S. wallisii as a first step of MMCT in aroid interspecific or intergeneric breeding.

The sprout inhibitors chlorpropham and 1,4-dimethylnaphthalene elicit different transcriptional profiles and do not suppress growth through a prolongation of the dormant state.

Chlorpropham (CIPC) and 1,4-dimethylnapthalene (DMN) are used to control postharvest sprouting of potato tubers. CIPC alters microtubule structure and function resulting in inhibition of cell division. The mechanism of action of DMN is unknown but, because it is a natural product found in potato tubers, there is speculation that it inhibits sprout growth by prolonging the dormant state. To address this issue, the effects of CIPC and DMN on abscisic acid (ABA) content and gene expression in potato tuber meristems were determined and compared to those found in dormant and non-dormant meristems. Dormancy progression was accompanied by a dramatic decline in ABA content and the ABA levels in meristems isolated from CIPC- and DMN- treated tubers were identical to the levels found in nondormant meristems demonstrating that sprout repression is not a function of elevated ABA. Evaluation of transcriptional profiles using cDNA microarrays demonstrated that there were similarities between CIPC- and DMN- treated tuber tissues particularly in transcripts that encode phosphatases and proteins associated with oxygen-related metabolism. Despite these similarities, there were significant differences in transcript profiles derived from treatment with either CIPC or DMN and the dormant state. These results suggested the mechanisms-of -action of DMN and CIPC are distinct and not due to a prolongation of the normal dormant condition.

Level and fate of chlorpropham in potatoes during storage and processing.

Chlorpropham (isopropyl 3-chlorocarbanilate) is a pesticide used to control sprouting of potatoes during long-term storage. The objective of the present study was to establish the total chlorpropham residue balance (residues in unwashed and washed whole tubers, peeled tubers, peels, boiled and pureed tubers, and washing and cooking waters) for two potato varieties after uniform application as a function of storage time under different typical storage conditions (in a basement, storehouse, and refrigerator) in Lithuania. Chlorpropham concentration on washed and unwashed tubers decreased from approximately 15 mg kg(- 1) after storage for 28 d to approximately 9 mg kg(- 1) after storage for 85 d. Peel concentrations decreased from approximately 50 mg kg(- 1) at 5 d after treatment to approximately 20 mg kg(- 1) at 85 d after treatment. The average concentration in the two varieties of peeled tubers in the three storage facilities was 1 mg kg(- 1). Chlorpropham concentrations in the wash water decreased from 3.5 to 1.0 mg L(- 1) when the storage time increased from 28 to 85 d. The cooking water concentration similarly decreased, from > 0.2 mg L(- 1) at 28 d after treatment to > 0.1 mg L(- 1) at 85 d after treatment. The results of this study show that chlorpropham concentrations in tubers under these typical conditions were below the recently revised levels that are acceptable for residues in whole potatoes (30 mg kg(- 1)) and peels (40 mg kg(- 1)). Also, despite fluctuating conditions during storage, chlorpropham treated tubers did not sprout, as compared to untreated tubers, which sprouted.

Rapid determination of aniline metabolites of chlorpropham in potatoes by micellar electrokinetic chromatography using negative-charged mixed micelles and laser-induced fluorescence detection.

A rapid, reliable method has been developed for the multi-residue analysis of aniline metabolites of chlorpropham in potato samples. The method involves the precolumn derivatization of aniline metabolites with 5-(4,6-dichloro-s-triazin-2-ylamino) fluorescein (DTAF) and their subsequent separation and determination by micellar electrokinetic capillary chromatography with laser-induced fluorescence detection (MEKC-LIF). The optimum procedure includes a derivatization step of the aniline metabolites (3-chloroaniline, 3-chloro-4-hydroxyaniline and 3-chloro-4-methoxyaniline) at 40 degrees C for 40 min and a 5-fold dilution prior to MEKC analysis, which is conducted within about 7 min using negative-charged mixed micelles (SDS/Triton X-100) in the running buffer. Under these conditions, the DTAF-anilines were readily detected at 0.3-3.1 microg/L level with a precision of 4.8-6.4%. These results indicate that negative-charged mixed surfactant MEKC-LIF is useful as a selective, rapid, and sensitive tool for the determination of these anilines and surpasses other electrophoretic alternatives based on the use of fluorescein-isothiocyanate (FITC) as label reagent. Finally, the potato matrix showed no significant effects on the derivatization and determination of these analytes, since the analytical figures of merit for the real samples were similar to those obtained in aqueous solutions, and the average recovery at fortification levels of 10-250 microg/kg was over 97%.

The heterogeneity of sprout inhibitor application with chlorpropham.

The CIPC or chlorpropham is used on potatoes as "sprouting inhibitor". These lasts years, some set of belgian potatoes treated by CIPC exceeded the Maximum Residue Limit (MRL), fixed at 5 mg/kg. The heterogeneity of sprout inhibitor application would be one of the causes of over-dosage. In order to estimate the distribution of CIPC between potatoes, according to the formulation used (DP, EC and HN), a research project financed by "le fond budgétaire des matières premieres" has been set up. In order to evaluate the distribution of CIPC into the pile of potatoes, the efficiency of the different formulations as well as the residues caused by their application on tubers, some tests have been placed in the storage hall. These tests consist in taking off some samples at different places into the pile of potatoes during storage and destocking. The content of CIPC is analysed by capillary gas chromatography with detection by mass spectrometry detection (GC-MS) or nitrogen specific detection (GC-NPD). To estimate the efficiency, the sprouting are observed every month and samples of potatoes are taken off on the top of the pile to control the evolution of CIPC content. Considering the results of the tests, the inhibitor treatment with the formulation DP+HN seems to be the more efficient. As for the residues, the formulation DP leads to a higher content of CIPC than HN, that has a very low concentration. After treatment, the quantity really applied on tubers depends on the formulation used. In fact, 50% of CIPC applied by DP formulation are found on tubers but hardly 10%, after gas application.

Study of potatoes' sprout inhibitor treatments with chlorprophame.

Studies carried out in 1999 by the University of Ghent showed that 36% of potatoes' samples contained Chlorprophame (CIPC) residues and that 7.9% of them exceeded the maximal limit of residues (RML), fixed at 5 ppm. The heterogeneity of sprout inhibitor application would be one of the causes of over-dosage. However, this heterogeneity would also cause under-dosages leading to problems when controlling the sprouting in potatoes stored over 6 degrees C. This study aims at determining some technical causes of the heterogeneity of CIPC sprout inhibitor treatments when storing potatoes. The study concerns two treatment techniques: dusting and spraying. To draw up an inventory of mechanical treatments in Belgium, a survey has been conducted among 28 farmers throughout Belgium. 35 samples have been taken at random in the different storage rooms to analyse the content of CIPC residue. In order to do so, a method of analysis: the gas chromatography in capillary phase with detection by mass spectrophotometry, has been developed. Tests have been carried out by changing several parameters such as the material, the product or the place in the storage line, in order to assess the CIPC application techniques. The survey made it possible to analyse qualitatively, from the declarations of farmers, the causes of heterogeneity linked to treatment techniques. An almost systematically over-dosage of the CIPC quantity has been noticed. However, out of the 35 samples analysed, only 2 had residue contents higher than the RML. The comparative analysis of the quantities applied and the residues contained in the samples made it possible to quantify the heterogeneity of the applications depending on the techniques. The tests carried out show in a general way that mechanical dusting, even though having a less constant flowrate than sprayers, leads to less important variation of the residue between samples. In testing conditions, the heterogeneity of the antigerminative treatment decreases when applied by means of a mechanical duster. In practise, these results are distorted by topical applications of CIPC. The combination of this practise with a too high heterogeneity of the treatment are to be avoided in order to have a good preservation and meet the residues standards.

Residue levels of chlorpropham in individual tubers and composite samples of postharvest-treated potatoes.

Chlorpropham, a herbicide and sprout suppressant, is used on stored potatoes to prolong the storage period without deterioration of produce quality. Data for residue concentrations on an individual tuber basis are required by WHO for the estimation of the variability factor. In this study, the levels of chlorpropham in individual tubers and in composite samples were determined. The distribution of chlorpropham between the peel and the tuber flesh was examined, and the fate during the cooking process (washing, boiling, frying) was studied. The concentrations in individual tubers ranged from 1.8 to 7.6 mg/kg 10 days postapplication (mean 3.8 mg/kg, RSD 39%), from 0.7 to 4.0 mg/kg 28 days postapplication (mean 2.9 mg/kg, RSD 28%), and from 0.8 to 3.8 mg/kg 65 days postapplication (mean 2.2 mg/kg, RSD 48%). The calculated residues in composite samples 10 days postapplication ranged from 4.3 to 6.1 mg/kg (mean 4.9 mg/kg, RSD 20%). Those in samples taken 28 days postapplication ranged from 3.1 to 4.2 mg/kg (mean 3.8 mg/kg, RSD 15%). The concentrations determined in composite samples of whole tubers 65 days postapplication ranged between 2.6 and 3.2 (mean 2.9 mg/kg, RSD 11%). Peeling removed 91-98% of the total residue; washing reduced residues by 33-47%. Detectable residues were found in boiled potatoes and the boiling water, and in french fries and the frying oil. Monitoring data on commercial prefried frozen french fries are reported.

Some applications of solid phase micro extraction (SPME) in the analysis of pesticide residues in food.

The Solid Phase Micro-Extraction (SPME) technique was applied to analyze chlorpropham in potatoes and amitraz in honey. The homogenized sample, suspended in water and stirred, was extracted with a 100 microns thick polidimetylsiloxane fiber and desorbed into the injection port of a gas chromatograph/mass-spectrometer (quadrupole) operating in single ion monitoring. Sensitivities down to 0.01 mg/Kg and linear responses in the range of 0.01-0.1 mg/Kg were obtained. The results of SPME pesticide residue analysis in potatoes corresponded to those obtained with a traditional multiresidue method.

The carry-through of residues of thiabendazole, tecnazene and chlorpropham from potatoes following manufacture into potato crisps and jacket potato crisps.

Potatoes, commercially treated with thiabendazole, tecnazene and chlorpropham, were processed into potato crisps and jacket potato crisps at a crisp factory using standard manufacturing conditions. A multi-residue method based on gas chromatography with mass spectrometric detection was developed and used to determine pesticide residue levels in the potatoes and potato crisps. Results showed that the residues of all three pesticides were significantly reduced to less than 2% and less than 10% of the maximum theoretical residue carry-through level for potato crisps and jacket potato crisps respectively.

A simple and quick gas chromatographic method for the determination of propham and chlorpropham in potatoes.

This study describes a gas chromatographic method for the quantitative determination of residual propham and chlorpropham in potatoes. Both herbicides are extracted from the foodstuff with methylene chloride. After centrifugation and concentration, propham and chlorpropham are quantitatively determined by gas chromatography thermionic detection using a fused silica capillary column CP Sil 5CB. 2-Chloraniline is used as internal standard. Recoveries of 100 +/- 15% and 99 +/- 10% have been obtained for propham and chlorpropham in blank samples spiked at the level of 0.5, 1.0 and 5.0 mg/kg. The absolute detection limit for both compounds is 1 ng corresponding to 0.1 mg/kg. Of the 161 samples of fresh potatoes analysed using this method, 136 contained residues of these herbicides and 18 of them (11%) exceeded the maximum tolerated value of 5 mg/kg.

Comparison of an HPTLC and an HPLC procedure for the determination of chlorpropham, propham and thiabendazole residues in potatoes.

HPTLC was used to check for residues of chlorpropham, propham (sprout inhibitors) and thiabendazole (fungicide) applied to potatoes. The method used gave good precision and analytical sensitivity. The sample preparation method developed by the authors was simple and gave good recovery and selectivity as far as other components of the matrix were concerned. Comparison with liquid chromatography confirmed the validity of the results.