Chitosan coated calcium alginate beads for covalent immobilization of acrylamidase: Process parameters and removal of acrylamide from coffee.

This study reports on removal of acrylamide from roasted coffee by acrylamidase from Cupriavidus oxalaticus ICTDB921. Chitosan coated calcium alginate beads were functionalized with citric acid as nontoxic cross linker and activated by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) (1.66:1 w/w) for covalent immobilization of acrylamidase. The optimum beads were obtained using 5% sodium alginate, 1.5% chitosan, and 0.6 mol/L citric acid. The beads prepared at each step were characterized by FTIR and SEM. Coating of chitosan matrix on calcium alginate beads enhanced the mechanical stability over that of calcium alginate and/or chitosan. The immobilized acrylamidase showed optimum pH/temperature of 8.5/65 °C, improved pH/thermal/shelf stability, and retained 80% activity after four cycles. Haldane model could describe the degradation kinetics of acrylamide in batch study. In packed bed column, a bed height, feed flow rate and inlet acrylamide concentration of 20 cm, 1 mL/min, and 100 mg/L gave best results.

Determination of acrylamide in brewed coffee by dispersive liquid-liquid microextraction (DLLME) and ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS).

This work has proposed the application of optimized dispersive liquid-liquid microextraction (DLLME) in order to extract acrylamide from brewed coffee samples for its subsequent determination by ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). DLLME achieved superior results employing 300 µL of brewed coffee, 100 µL of dichloromethane, 400 µL of acetonitrile and without sodium chloride addition. Quantitative analyses were carried out by the standard addition method, and the limits of detection and quantification were 0.9 and 3.0 µg L-1, respectively. Recoveries ranged from 97 to 106%, and the intra- and inter-assay precisions ranged from 6 to 9%. The proposed analytical method was applied to seventeen brewed coffee samples prepared in a filter coffee maker, and acrylamide amounts varied from 10.5 to 28.5 µg L-1. Therefore, the suggested DLLME-UPLC-MS/MS method is promising for routine analysis in order to guarantee the quality control of acrylamide in brewed coffee.

Single-drop microextraction combined with gas chromatography-electron capture detection for the determination of acrylamide in food samples.

A single-drop microextraction method followed by gas chromatography-electron capture detection was developed to determine acrylamide in food samples. Acrylamide was extracted by water and derivatized by hydrobromic acid in the presence of ammonium peroxydisulfate. The derivatization was carried out at 45 °C in 15 min using 46 µL of hydrobromic acid and 98 mg of ammonium peroxydisulfate. A 3.0-mL volume of the derivatized analyte was extracted using a 1.0-µL n-octanol droplet hanging from the needle tip of a GC microsyringe. After extraction, the extract was injected into the gas chromatograph. The influence of experimental parameters effective on derivatization reaction yield and extraction performance was studied. The limit of detection and quantification, relative standard deviation and linearity of the method were 0.60 µg/L, 2.0 µg/L, <6.0%, and 2.0-100.0 µg/L, respectively. The method was utilized to determine acrylamide in three food samples (i.e., bread, potato chips and cookie).

An electrochemical biosensor based on hemoglobin-oligonucleotides-modified electrode for detection of acrylamide in potato fries.

Acrylamide a neurotoxin and strong carcinogen, is found in various thermally processed foods. In this study, an electrochemical sensor for detection of acrylamide using double stranded DNA (dsDNA)/Hemoglobin (Hb)-modified screen printed gold electrode (SPGE) was designed. The immobilization of ssDNA1-SH on the surface of SPGE was confirmed by cyclic voltammetry, and the interaction between ssDNA2-NH2 and Hb with the ratio 1:1 was characterized by agarose gel. The excellent response of the designed biosensor towards acrylamide due to acrylamide and Hb adducts and change of reduction/oxidation process of Hb-Fe(III)/Hb-Fe(II) was determined by square wave voltammetry (SWV). The biosensor showed the optimum response at pH 8.0. The linear working range for acrylamide was from 2.0 × 10-6 to 5.0 × 10-2 M with a detection limit of 1.58 × 10-7 M. The biosensor was suitable for direct determination of acrylamide in water extracted of potato fries and displayed good reproductivity and high stability.

Biochemical characterization of a novel L-asparaginase from Paenibacillus barengoltzii being suitable for acrylamide reduction in potato chips and mooncakes.

A novel L-asparaginase gene (PbAsnase) from Paenibaeillus barengoltzii CAU904 was cloned and expressed in Escherichia coli. The L-asparaginase gene was 1011bp encoding 336 amino acids. Multiple sequence alignment of PbAsnase with other known L-asparaginases revealed that the enzyme showed high similarities with some Rhizobial-type L-asparaginases, sharing the highest identity of 32% with a characterized L-asparaginase from Rhizobium etli CFN 42, suggesting that it should be a novel L-asparaginase. The recombinant L-asparaginase (PbAsnase) was purified to homogeneity and biochemically characterized. The purified enzyme was optimally active at pH 8.5 and 45°C, respectively. It was stable within pH 5.5-10.0 and at temperatures below 55°C. PbAsnase exhibited strict substrate specificity towards L-asparagine (35.2U/mg), with Km and Vmax values of 3.6mM and 162.2µmol/min/mg, respectively, but displayed trace activity towards L-glutamine. Moreover, the application potential of PbAsnase on acrylamide migration in potato chips and mooncakes was evaluated. The pretreatment by PbAsnase significantly decreased the acrylamide contents in potato chips and mooncakes by 86% and 52%, respectively. The unique properties of PbAsnase may make it a good candidate in industries, especially in food safety.

Matrix-compatible sorbent coatings based on structurally-tuned polymeric ionic liquids for the determination of acrylamide in brewed coffee and coffee powder using solid-phase microextraction.

Nine crosslinked polymeric ionic liquid (PIL)-based SPME sorbent coatings were designed and screened in this study for the trace level determination of acrylamide in brewed coffee and coffee powder using gas chromatography-mass spectrometry (GC-MS). The structure of the ionic liquid (IL) monomer was tailored by introducing different functional groups to the cation and the nature of the IL crosslinker was designed by altering both the structure of the cation as well as counteranions. The extraction efficiency of the new PIL coatings towards acrylamide was investigated and compared to a previously reported PIL sorbent coating. All PIL fibers exhibited excellent analytical precision and linearity. The PIL fiber coating consisting of 50% 1,12-di(3-vinylbenzylbenzimidazolium)dodecane dibis[(trifluoromethyl)sulfonyl]imide as IL crosslinker in 1-vinyl-3-(10-hydroxydecyl)imidazolium bis[(trifluoromethyl)sulfonyl]imide IL monomer resulted in a limit of quantitation of 0.5µgL(-1) with in-solution SPME sampling. The hydroxyl moiety appended to the IL cation was observed to significantly increase the sensitivity of the PIL coating toward acrylamide. The quantitation of acrylamide in brewed coffee and coffee powder was performed using the different PIL-based fibers by the method of standard addition after a quenching reaction using ninhydrin to inhibit the formation of interfering acrylamide in the GC inlet, mainly by asparagine thermal degradation. Excellent repeatability with relative standard deviations below 10% were obtained on the real coffee samples and the structure of the coatings appeared intact by scanning electron microscopy after coffee sampling proving the matrix-compatibility of the PIL sorbent coatings.

Direct determination of acrylamide in potato chips by using headspace solid-phase microextraction coupled with gas chromatography-flame ionization detection.

Acrylamide is a potentially toxic and carcinogenic substance present in many high-consumption foods. Recently, this matter has been placed in category of "reasonably anticipated to be a human carcinogen" by National Toxicology Program (NTP). Therefore, simple and cost-effective determination of acrylamide in food samples has attracted intense interest. The most reported techniques for this purpose are GC-MS and LC-MS, which are very expensive and available in few laboratories. In this research, for the first time, a rapid, easy and low-cost method is introduced for sensitive and precise determination of acrylamide in foodstuffs, using gas chromatography-flame ionization detection (GC-FID) system after its direct trapping in the upper atmosphere of samples by headspace solid-phase microextraction (HS-SPME). The effects of main experimental variables were studied and the optimized parameters were obtained as the type of fiber, carboxen/divinylbenzene/polydimethylsiloxane (CAR/DVB/PDMS); extraction time, 30 min; extraction temperature, 60°C; moisture content, 10 µL water per 1g of sample; desorption time, 2 min; and desorption temperature, 230°C. The linear calibration graph was obtained in the range of 0.77-50 µg g(-1), with regression coefficient of 0.998. The detection and quantification limits of the proposed method were 0.22 and 0.77 µg g(-1), respectively. The recoveries, for different food samples, were 79.6-95.7%. The repeatability of measurements, expressed as relative standard deviation (RSD), were found to be 4.1-8.0% (n=9). The proposed HS-SPME-GC-FID method was successfully carried out for quantifying of trace levels of acrylamide in some processed food products (chips and French fries), sold in open local markets.