Reduce Acrylamide Formation Research Articles

Potassium Hydroxide Extraction of Polyphenols from Olive Leaves: Effect on Color and Acrylamide Formation in Black Ripe Olives

Olive leaves are generated in large quantities in olive oil and table olive factories. This waste is currently used for multiple purposes, one of them being the extraction of bioactive substances, in particular phenolic compounds. The aims of this study were (i) to obtain a new polyphenolic extract from potassium hydroxide (KOH) -treated olive leaves; and (ii) to reduce acrylamide formation in black olives by using this extract. The results showed that olive leaves and leafless branches of the Manzanilla cultivar treated with 10 g/L KOH provide a solution that, concentrated under vacuum, had >6000 mg/L hydroxytyrosol and >2000 mg/L of hydroxytyrosol 4-glucoside. Moreover, the residual material generated after the treatment with KOH could be used for agronomic purposes, due to its high potassium content. The employment of this non-bitter extract during the darkening step of black ripe olive processing then resulted in darker fruits with higher potassium content. Likewise, the addition of the extract into the packing brine reduced the acrylamide formation by up to 32%, although this effect was batch-dependent. KOH olive extract could be useful for the reduction in acrylamide in black ripe olives along with the enrichment of this product in phenolic compounds and potassium.

Engineering Tk1656, a highly active l-asparaginase from Thermococcus kodakarensis, for enhanced activity and stability

l-Asparaginases catalyze the hydrolysis of l-asparagine to l-aspartic acid and ammonia. These enzymes have potential applications in therapeutics and food industry. Tk1656, a highly active and thermostable l-asparaginase from Thermococcus kodakarensis, has been proved effective in selective killing of acute lymphocytic leukemia cells and in reducing acrylamide formation in baked and fried foods. However, it displayed <5 % activity under physiological conditions compared to the optimal activity at 85 °C and pH 9.5. We have attempted engineering of this valuable enzyme to improve the characteristics required for therapeutic and industrial applications. Based on the literature and crystal structure of Tk1656, nine specific mutant variants were designed, produced in Escherichia coli, and the purified mutant enzymes were compared with the wild-type. One of the mutants, K299L, displayed >20 % increase in activity at 85 °C. H158S substitution resulted in >5 °C increase in the optimal temperature. Similarly, a mesophilic-like mutation L56D, resulted in >5-fold increase in activity at pH 7.0 and 37 °C compared to that of the wild-type enzyme. The substrate specificity of the mutant variants remained unchanged. These results demonstrate that L56D and K299L variants of Tk1656 are the potent enzymes for therapeutics and acrylamide mitigation applications, respectively.

Application of Cations, Acids, and Antioxidants on Mitigating Acrylamide Formation in Food with a Special Focus on Potato and Cereal Based Foods - A Review

Acrylamide is a toxic substance that formsin food during high-temperature processing methods such as deep-frying, baking, and roasting, through the Maillard reaction or the acrolein pathway. Since acrylamide has been identified for its carcinogenicity, genotoxicity, and neurotoxicity, global food authorities have established benchmark levels for different food categories. Consequently, researchers have proposed various mitigation strategies, focusing on agronomical, chemical, physical, and microbial approaches. However, most of these approaches are associated with high cost, technology, extended time, and poor sensory and nutritional quality. In this context, application of various additives such as metal cations, organic and inorganic acids, and antioxidants in optimum amounts have been effective in reducing acrylamide formation. Additives contribute to reducing acrylamide either by interacting with its precursors and/or intermediate products or by breaking down acrylamide into other non-toxic substances. Since cations can form thermostable asparagine/matrix intermediates through chelation reaction between asparagine and metal cations, asparagine will be unavailable to react with carbonyl compounds to form the Schiff base. Lower pH conditions inhibit the acrylamide formation by blocking the protonation of the α-amino group of asparagine and reducing any possibility for nucleophilic addition reactions with carbonyl groups. Toxicity associated with Maillard products can be reduced by using antioxidants in the frying oil or the food. Polyphenols are capable of reducing the acrylamide formation by directly trapping acrylamide at elevated temperature. It is essential to identify the most appropriate additive, its concentration and the optimum process conditions when applying additives to get an optimal acrylamide inhibition. This review article highlights the effect of various additive applications, their effectiveness and possible future advances in food to control acrylamide formation within the accepted level.Keywords: Acrylamide, Antioxidant, Carcinogenicity, Deep-frying, Maillard reaction

Reducing Acrylamide Formation Potential by Targeting Free Asparagine Accumulation in Seeds.

Acrylamide is a probable carcinogen in humans and is formed when reducing sugars react with free asparagine (Asn) during thermal processing of food. Although breeding for low reducing sugars worked well in potatoes, it is less successful in cereals. However, reducing free Asn in cereals has great potential for reducing acrylamide formation, despite the role that Asn plays in nitrogen transport and amino acid biosynthesis. In this perspective, we summarize the efforts aimed at reducing free Asn in cereal grains and discuss the potentials and challenges associated with targeting this essential amino acid, especially in a seed-specific manner.

Effect of ultrasound and additives treatment as mitigation strategies to reduce acrylamide formation in potato crisps on industrial scale

The aim of this work was to examine the applicability on large scale of additives and ultrasound treatments during soaking of potatoes before frying to mitigate the formation of acrylamide in potato crisps. Calcium chloride and citric acid were applied at laboratory scale in various concentrations and orders during washing before frying, to establish optimum conditions which were scaled up to pilot plant. Up to 91.0% reduction in acrylamide was obtained at laboratory scale. Both concentration and order of additives influenced the extent of the mitigation observed, with a higher concentration of additive in the second wash being beneficial. When upscaled to factory pilot plant, the reduction observed was not consistent across the three trials, with a 33.4% reduction in the first trial but no significant reduction in following studies. A 2-min ultrasound treatment was applied in two trials to test various powers and amplitudes, and washing combinations respectively. Up to 67.1% of acrylamide reduction was recorded after 2 min of ultrasound treatment in the cold wash followed by hot wash; however, ultrasound treatment was not effective in reducing acrylamide or its precursors when solely applied or when followed by cold wash under the tested conditions of duration and power.

The efficacy and lipid degradation properties resulting from corncob biochar treatment for acrylamide reduction in reused palm oil

Acrylamide is a carcinogenic compound commonly found in deep fried food. Given the health adverse effects of acrylamide, innovative techniques that can reduce acrylamide formation during frying is desirable while maintaining oil quality. This study examined how used palm oil treated with corncob biochar changed the amount of acrylamide and the composition of lipids. The biochar was mixed with used oil (ratio 1:10 (w/w)) for 15 h and the chemical properties of the oil were determined. The properties of biochar treated oil were compared with fresh oil, which formed the control, and used oil prior to biochar treatment. The results of oxidation (PV, TBARs) were significantly lower when the used oil was treated before being fried again. The use of biochar did not alter the fatty acid profile of the oil. Acrylamide content in fried potatoes was reduced by half compared to 2nd used oil (2 mg/100g) and 2nd used oil treated with biochar (0.8 mg/100g). This preliminary study shows that biochar made from agricultural waste has the potential to be used as an oil recycling material for a more sustainable alternative approach. Moreover, the method is applicable to industrial contexts and the material is safe for the environment.

Psyllium husk gel used as an alternative and more sustainable scalding technology for wheat bread quality improvement and acrylamide reduction.

This study aimed at evaluating the influence of different amounts (5, 10, 15, 20, and 25%) of psyllium husk gel (PHG) on wheat bread (WB) characteristics - chiefly, overall acceptability (OA), porosity, specific volume (v), mass loss after baking (ML), shape retention coefficient, crust and crumb color coordinates, bread crumb hardness during storage, saccharides content, and acrylamide (AA) concentration. PHG was prepared by mixing 100 g of psyllium husk powder with 800 mL of warm water. It was established that the amount of psyllium husk gel is a significant factor in dough redness (a*) (p < 0.001). A moderate positive correlation (r) was found between acrylamide content in wheat bread and maltose concentration in dough (r = 0.567). The psyllium husk gel increased the overall acceptability and specific volume of wheat bread. Wheat bread porosity showed a moderate positive correlation with mass loss after baking (r = 0.567) and a strong positive correlation with texture hardness (r = 0.664). Lower acrylamide content was obtained in wheat bread prepared with 5, 10, 15, 20, and 25% of psyllium husk gel (1.53, 2.34, 3.80, 2.69, and 3.62 times lower than the control wheat bread, respectively). Acrylamide content showed a strong positive correlation with the porosity of wheat bread (r = 0.672), with crust brightness (L*), and yellowness/blueness (b*) coordinates, as well as with crumb brightness, redness, and yellowness coordinates. Overall, psyllium husk gel hydrocolloids reduced acrylamide formation in wheat bread and can be recommended for the quality improvement of wheat bread.

Enhancing biotechnological applications of l-asparaginase: Immobilization on amino-epoxy-agarose for improved catalytic efficiency and stability

l-asparaginase has been shown to be effective in reducing acrylamide formation in food and in treating certain cancers, such as acute lymphoblastic leukemia. This study aimed to improve the application of l-asparaginase in these areas, through its immobilization in two distinct supports: epoxy-agarose and amino-epoxy-agarose. Different immobilization conditions were investigated, including immobilization pH and buffer concentration. The best activity results were obtained employing amino-epoxy-agarose, being the 0.1 M sodium carbonate buffer pH 10 the conditions that offered the highest immobilization yield (76%), immobilization efficiency (89.8%) and relative activity (68.2%). Immobilization of l-asparaginase in amino-epoxy-agarose improved its catalytic efficiency and stability, including at pH 4 and 60 °C. All immobilized biocatalysts preserved more than 70% of their activity after 7 cycles of reuse, and maintained about 82% of their activity after 60 days of storage at 4°C. These results show the efficacy of immobilized biocatalysts in the preservation of enzymatic activity and highlight their potential for biotechnological applications.

Acrylamide and 5-Hydroxymethylfurfural in Synthetic Sugar Cane Syrup: Mitigation by Additives.

The ability of additives to reduce the formation of acrylamide in simulated sugar cane syrups was investigated. Organic acids, B vitamins, and inorganic salts were added individually and in combination to simulated thickened cane juice, and the mixtures were heated at 120 °C for 30 min. Calcium chloride (1%), citric acid (0.1%), and vitamin B3 (0.1%) were the most effective individual additives from each chemical family. The effects of CaCl2 (0-1%), citric acid (0-0.125%), and vitamin B3 (0-0.1125%), when added in combination, on the concentrations of acrylamide and hydroxymethylfurfural (HMF) were studied using a Box-Behnken design. Combinations of all three additives lowered the acrylamide production, but only the combination of citric acid and vitamin B3 had a significant synergistic effect. However, all these additives stimulated the production of HMF, and no significant interactive effect between pairs of additives on HMF production was observed. Calcium chloride stimulated the formation of HMF most strongly. These results indicate that certain combinations of these additives effectively reduce acrylamide formation, but they also lead to an increase in the formation of HMF in sugar syrup.

Vacuum impregnation pre-treatment: A novel method for incorporating mono- and divalent cations into potato strips to reduce the acrylamide formation in French fries

Abstract The effect of different vacuum impregnation (VI) pre-treatments of potato strips [in water (control), NaCl, KCl or CaCl2 (0.1 M) for 10 and 15 min] on acrylamide formation in French fries after deep frying in sunflower oil at 175°C for 5 min was determined. The acrylamide content of fries prepared with potato strips pre-treated by VI in distilled water decreased from 0.82 to 0.24 µg/g when time increased to 15 min. Moreover, the acrylamide content of fries from 10 min-KCl pre-treatment (0.35 µg/g) decreased significantly to 0.04 µg/g when time was 15 min (p &lt; 0.05). In comparison to control (10 min), the VI pre-treatment in KCl and NaCl for 15 min reduced the relative content of acrylamide by 95 and 91%, respectively, and VI in distilled water was able to decrease this value by 71%. Results indicated that monovalent cations (K and Na) were more effective in reducing acrylamide formation than divalent cation (Ca2+) when the VI pre-treatment time increased to 15 min. In conclusion, VI as a pre-treatment for mono- and divalent cations in fried potato production could be a useful technique in industrial applications to reduce acrylamide formation in fries in a short time.

Crickets (Acheta domesticus) as Wheat Bread Ingredient: Influence on Bread Quality and Safety Characteristics.

The aim of this study was to assess respondents' opinions on the choice of edible insects as a food, and to evaluate the influence of cricket flour (ECF) (10, 20, 30%) on the quality of wheat bread (WB). Whereas ECF is an additional source of acrylamide precursors, in order to reduce acrylamide formation in WB, fermentation of ECF with Lactiplantibacillus plantarum-No.122 was applied. It was established that 70.7% of the respondents had never eaten insects and more than 30% would not choose them. However, ECF was suitable substrate for fermentation (lactobacilli count 8.24 log10CFU/g, pH-4.26). In addition, fermentation reduced the total biogenic amines content in ECF (by 13.1%). The highest specific volume showed WB, prepared with fermented ECF (10, 20, 30%). All the tested WB showed similar overall acceptability (on average, 7.9 points). However, the highest intensity of emotion "happy" was induced by the WB, prepared with fermented ECF. Most of the WB with non-treated and fermented ECF showed higher acrylamide concentration (except WB with 10% of fermented ECF), in comparison with the control. Finally, fermentation is recommended for ECF inclusion in the main WB formula because fermentation improves not only quality but also reduces acrylamide concentration in WB.

Molecular Characterization of a Stable and Robust L-Asparaginase from Pseudomonas sp. PCH199: Evaluation of Cytotoxicity and Acrylamide Mitigation Potential

L-asparaginase is an important industrial enzyme widely used to treat acute lymphoblastic leukemia (ALL) and to reduce acrylamide formation in food products. In the current study, a stable and robust L-asparaginase from Pseudomonas sp. PCH199, with a high affinity for L-asparagine, was cloned and expressed in Escherichia coli BL21(DE3). Recombinant L-asparaginase (Pg-ASNase II) was purified with a monomer size of 37.0 kDa and a native size of 148.0 kDa. During characterization, Pg-ASNase II exhibited 75.8 ± 3.84 U/mg specific activities in 50.0 mM Tris-HCl buffer (pH 8.5) at 50 °C. However, it retained 80 and 70% enzyme activity at 37 °C and 50 °C after 60 min, respectively. The half-life and kd values were 625.15 min and 1.10 × 10−3 min−1 at 37 °C. The kinetic constant Km, Vmax, kcat, and kcat/Km values were 0.57 mM, 71.42 U/mg, 43.34 s−1, and 77.90 ± 9.81 s−1 mM−1 for L-asparagine, respectively. In addition, the enzyme has shown stability in the presence of most metal ions and protein-modifying agents. Pg-ASNase II was cytotoxic towards the MCF-7 cell line (breast cancer) with an estimated IC50 value of 0.169 U/mL in 24 h. Further, Pg-ASNase II treatment led to a 70% acrylamide reduction in baked foods. These findings suggest the potential of Pg-ASNase II in therapeutics and the food industry.

Addition of by-Products and Flavored Hydrocolloids Filling in Californian-Style Black Olive to Reduce Acrylamide Formation

Oxidized black table olives are frequently consumed in the Mediterranean diet. To improve food quality, the use of by-products is an emergent strategy that should be more studied. With a better understanding of the use of by-products, healthier food with the highest possible quality could be obtained, increasing its added value. Different by-products at different concentrations (1:10 and 1:100) and phenol-rich aromatized hydrocolloids were added to Californian-style black olive in order to evaluate its effect in acrylamide. In general, the addition of by-products before the sterilization process resulted in a significant reduction of the acrylamide content (10–60%). The highest inhibition was obtained with the phenolic extract at a 1:10 dilution. Furthermore, flavored olives stuffed with higher concentrations caused a greater reduction in acrylamide content. The ’Garlic’ and ‘Thyme’ flavors showed the greatest reduction of this toxic substance. The tasting panel considered that olives with a 1:10 dilution and olives filled with flavored hydrocolloids at low concentrations had a pleasant odor and positive sensory attributes. Orange peel was the recommended by-product to mitigate the effects of acrylamide since it obtained the highest score both in acrylamide inhibition and in the taste panel. As for the hydrocolloids, ‘Thyme’ showed the greatest inhibition effect but did not perform well in the taste panel, with similar effects to ‘Oregano’ and ‘Garlic.’ The current research attempts to underline the use of local by-products to obtain additives rich in antioxidant activity that could enhance oxidized black table olives, becoming a healthier, safer and better-quality products. Moreover, this connects with the idea of moving from a linear economy to a circular economy, obtaining a product of high economic value from a by-product.

Effectiveness of asparaginase on reducing acrylamide formation in bakery products according to their dough type and properties

Effect of asparaginase was tested in different biscuit and cracker doughs, and wafer batter by changing processing conditions such as enzyme dosage, dough resting time and temperature, mixing speed and time, and mixing procedure of the recipe components. Acrylamide reductions achieved were 96, 80, 54% in rotary cut biscuits, crackers and wire cut cookies, respectively due to high water activity of their dough. Asparaginase did not affect surface color or spread ratio. There was a positive correlation between asparagine content and acrylamide formation except for rotary molded biscuit doughs. No significant decrease was found in rotary molded biscuits because of low water (water activity of 0.70) and high fat content. This indicated that water activity of dough is an important factor for the effectiveness of asparaginase treatment. The results suggested that water activity value exceeding 0.75 is needed in the dough to effectively reduce asparagine, so acrylamide in bakery products.

Formation and mitigation of acrylamide in oven baked vegetable fries

Investigation into oven baked sweet potato and carrot fries at various temperatures and times demonstrated the in situ formation of acrylamide in an exponential manner. High levels of acrylamide were found in these food items: up to 327 µg/kg for sweet potato baked at 190 °C for 14 min, and 99 µg/kg for carrot baked at 190 °C for 13 min. Risk assessment via Margin of Exposures estimation showed that consumption of these fries might pose adverse health effects to consumers from toddlers to adults, especially when the fries were prepared at high temperatures above 175 °C and for a long time. Raw ingredient blanching and immersion in acetic acid prior to preparation have been proven to greatly reduce acrylamide formation, up to 99%. It is recommendable to apply these techniques either at industrial or domestic cooking scales to ensure minimal health risk from dietary exposure to acrylamide.

ACRYLAMIDE IN DEEP-FRYING VERSUS AIR FRYING FOODS AND MITIGATION TREATMENTS: A REVIEW

Frying can contribute to intensifying oxidative reactions and the formation of acrylamide (AA), which is a compound derived from the Maillard Reaction, potentially toxic. Even so, fried foods are one of the most consumed foods worldwide. Innovative technologies such as air frying (AF) have been developed to this end. Thus, the objectives of this review were to systematize data on AA content in foods, compare the AF technique with immersion frying (FRIT), and assess the efficacy of pretreatments to reduce AA in foods exposed to AF. AF showed an average percentage reduction of AA content and colorimetric indicator a* of 41.4% and 58.7%, respectively. Asparaginase and plant extracts were effective in reducing AA formation, with mean reduction of 20.94% and 67.01%, respectively. AF proved to be a potentially efficient method for reducing AA in foods, and the use of pretreatments proved to be effective in reducing its formation even further.

A Novel L-Asparaginase from Hyperthermophilic Archaeon Thermococcus sibiricus: Heterologous Expression and Characterization for Biotechnology Application.

L-asparaginase (L-ASNase) is a vital enzyme with a broad range of applications in medicine and food industry. Drawbacks of current commercial L-ASNases stimulate the search for better-producing sources of the enzyme, and extremophiles are especially attractive in this view. In this study, a novel L-asparaginase originating from the hyperthermophilic archaeon Thermococcus sibiricus (TsA) was expressed in Escherichia coli, purified and characterized. The enzyme is optimally active at 90 °C and pH 9.0 with a specific activity of 2164 U/mg towards L-asparagine. Kinetic parameters KM and Vmax for the enzyme are 2.8 mM and 1200 µM/min, respectively. TsA is stable in urea solutions 0–6 M and displays no significant changes of the activity in the presence of metal ions Ni2+, Cu2+, Mg2+, Zn2+ and Ca2+ and EDTA added in concentrations 1 and 10 mmol/L except for Fe3+. The enzyme retains 86% of its initial activity after 20 min incubation at 90 °C, which should be enough to reduce acrylamide formation in foods processed at elevated temperatures. TsA displays strong cytotoxic activity toward cancer cell lines K562, A549 and Sk-Br-3, while normal human fibroblasts WI-38 are almost unsensitive to it. The enzyme seems to be a promising candidate for further investigation and biotechnology application.

L-asparaginase: therapeutic use and applications in the food industry – a review

L-asparaginase (L-asnase) is an amino hydrolase that has been used in the last decades for leukemia treatment, which boosted scientific studies on production, purification and immobilization of this enzyme. More recently, L-asnase has called food industry attention because of its effect on acrylamide formation in fried and baked foods. Several studies have been carried out in order to evaluate the effect of L-asnase in reducing acrylamide formation in different food models. This review brings up an overview in L-asnase kinetic parameters from different sources, immobilization methods, its therapeutic use in leukemia treatment and food processing applications. This review also discusses acrylamide formation in fried and baked foods. Commercial L-asnase is produced by two microorganisms, Escherichia coli and Erwinia sp. However, studies using different microorganisms have shown the possibility of producing this enzyme from different sources, obtaining enzymes with interesting kinetic properties. Immobilization strategies have provided enzymes with greater activity and stability, which could contribute to maintain L-asnase activity in the body for longer periods. Researches applying L-asnase in food products have shown significant reduction in acrylamide production, above 90% in some cases. For this purpose, during enzyme application some variables must be taken into account, as enzyme dose, food matrix, pretreatment, processing time and temperature. Medical and food applications make L-asnase a multipurpose enzyme. Reducing prices, improving enzyme stability and reducing co-lateral effects in leukemia treatment are still challenges to overcome.

Immobilization of L-asparaginase on magnetic nanoparticles: Kinetics and functional characterization and applications

L-asparaginase shows great potential as a food enzyme to reduce acrylamide formation in fried and baked products. But for food applications, enzymes must be stable at high temperatures and have higher catalytic efficiency. These desirable characteristics are conferred by the immobilization of enzymes on a suitable matrix. The present study aimed to immobilize the L-asparaginase enzyme on magnetic nanoparticles to reduce acrylamide content in the food system. Immobilized preparations were characterized using SEM, TEM, FTIR, UV-spectrometry, and XRD diffraction analyses. These nanoparticles enhanced the thermal stability of the enzyme up to four-fold at 70 °C compared to the free enzyme. Kinetic parameters exhibited an increase in Vmax, Km, and catalytic efficiency by ~ 38% than the free counterpart. The immobilized preparations were reusable for up to five cycles. Moreover, their application in the pre-treatment coupled with blanching of potato chips led to a significant reduction (greater than 95%) of acrylamide formation.

Reduction of acrylamide formation in bread and fried potato products using probiotic microorganisms: a systematic review and dose–response meta-analysis

Reduction of acrylamide formation in bread and fried potato products using probiotic microorganisms: a systematic review and dose–response meta-analysis