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Food Research International (Ottawa,... Oct 2023Acrylamide (AA), as a food-borne toxicant, is created at some stages of thermal processing in the starchy food through Maillard reaction, fatty food via acrolein route,... (Review)
Review
Acrylamide (AA), as a food-borne toxicant, is created at some stages of thermal processing in the starchy food through Maillard reaction, fatty food via acrolein route, and proteinous food using free amino acids pathway. Maillard reaction obviously takes place in thermal-based products, being responsible for specific sensory attributes; AA formation, thereby, is unavoidable during the thermal processing. Additionally, AA can naturally occur in soil and water supply. In order to reduce the levels of acrylamide in cooked foods, mitigation techniques can be separated into three different types. Firstly, starting materials low in acrylamide precursors can be used to reduce the acrylamide in the final product. Secondly, process conditions may be modified in order to decrease the amount of acrylamide formation. Thirdly, post-process intervention could be used to reduce acrylamide. Conventional or emerging mitigation techniques might negatively influence the pleasant features of heated foods. The current study summarizes the effect of enzymatic reaction induced by asparaginase, glucose oxidase, acrylamidase, phytase, amylase, and protease to possibly inhibit AA formation or progressively hydrolyze formed AA. Not only enzyme-assisted AA reduction could dramatically maintain bio-active compounds, but also no damaging impact has been reported on the sensorial and rheological properties of the final heated products. The enzyme engineering can be applied to ameliorate enzyme functionality through altering the amino acid sequence like site-specific mutagenesis and directed evolution, chemical modifications by covalent conjugation of L-asparaginase onto soluble/insoluble biocompatible polymers and immobilization. Moreover, it would be possible to improve the enzyme's physical, chemical, and thermal stability, recyclability and prevent enzyme overuse by applying engineered ones. In spite of enzymes' cost-effective and eco-friendly, promoting their large-scale usages for AA reduction in food application and AA bioremediation in wastewater and soil resources.
Topics: Asparaginase; 6-Phytase; Acrolein; Acrylamide; Amino Acid Sequence
PubMed: 37689930
DOI: 10.1016/j.foodres.2023.113177 -
Preparative Biochemistry & Biotechnology 2022L-asparaginases prevent the formation of acrylamide, a substance commonly found in foods subjected to heat and that also contains reducing sugars and L-asparagine. This...
L-asparaginases prevent the formation of acrylamide, a substance commonly found in foods subjected to heat and that also contains reducing sugars and L-asparagine. This work aimed to select a strain of spp. able to produce L-asparaginase and to optimize the fermentation parameters, the partial purification and biochemical characterization were also performed. The IOC 3999 was selected due to its greater enzymatic activity: 1443.57 U/mL of L-asparaginase after 48 h of fermentation. The optimized conditions allowed for an increase of 223% on the L-asparaginase production: 2.9% lactose, 2.9% L-asparagine and 0.7% hydrolyzed casein, 0.152% KHPO, 0.052% KCl and MgSO, 0.001% of CuNO.3HO, ZnSO.7HO and FeSO.7HO adjusted to pH 7.0; added a concentration of 5.05x10 spores/mL at 30 °C for 100 rpm. A purification factor of 2.11 was found and the molecular mass was estimated at 20.8 kDa. The enzyme showed optimum activity at 60 °C and pH 5 and stability at 50 °C for 1 h. The enzyme presented desirable biochemical characteristics, mainly the acid pH stability, indicating that the enzyme would work well in food matrices due to the closeness of pH, meaning that it could be a potential option for use in the food industry.
Topics: Asparaginase; Aspergillus oryzae; Culture Media; Enzyme Stability; Fermentation; Hot Temperature; Hydrogen-Ion Concentration; Hydrolysis
PubMed: 34110268
DOI: 10.1080/10826068.2021.1931881 -
Applied Microbiology and Biotechnology Jun 2021In the past decades, the production of biopharmaceuticals has gained high interest due to its great sensitivity, specificity, and lower risk of negative effects to... (Review)
Review
In the past decades, the production of biopharmaceuticals has gained high interest due to its great sensitivity, specificity, and lower risk of negative effects to patients. Biopharmaceuticals are mostly therapeutic recombinant proteins produced through biotechnological processes. In this context, L-asparaginase (L-asparagine amidohydrolase, L-ASNase (E.C. 3.5.1.1)) is a therapeutic enzyme that has been abundantly studied by researchers due to its antineoplastic properties. As a biopharmaceutical, L-ASNase has been used in the treatment of acute lymphoblastic leukemia (ALL), acute myeloblastic leukemia (AML), and other lymphoid malignancies, in combination with other drugs. Besides its application as a biopharmaceutical, this enzyme is widely used in food processing industries as an acrylamide mitigation agent and as a biosensor for the detection of L-asparagine in physiological fluids at nano-levels. The great demand for L-ASNase is supplied by recombinant enzymes from Escherichia coli and Erwinia chrysanthemi. However, production processes are associated to low yields and proteins associated to immunogenicity problems, which leads to the search for a better enzyme source. Considering the L-ASNase pharmacological and food importance, this review provides an overview of the current biotechnological developments in L-ASNase production and biochemical characterization aiming to improve the knowledge about its production. KEY POINTS: • Microbial enzyme applications as biopharmaceutical and in food industry • Biosynthesis process: from the microorganism to bioreactor technology • Enzyme activity and kinetic properties: crucial for the final application.
Topics: Antineoplastic Agents; Asparaginase; Asparagine; Biotechnology; Dickeya chrysanthemi; Escherichia coli; Humans; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Recombinant Proteins
PubMed: 34059941
DOI: 10.1007/s00253-021-11359-y -
Frontiers in Immunology 2024Asparaginase (ASNase) is a crucial part of acute leukemia treatment, but immune responses to the agent can reduce its effectiveness and increase the risk of relapse....
BACKGROUND
Asparaginase (ASNase) is a crucial part of acute leukemia treatment, but immune responses to the agent can reduce its effectiveness and increase the risk of relapse. Currently, no reliable and validated biomarker predicts ASNase-induced hypersensitivity reactions during therapy. We aimed to identify predictive biomarkers and determine immune cells responsible for anaphylaxis using a murine model of ASNase hypersensitivity.
METHODS
Our preclinical study uses a murine model to investigate predictive biomarkers of ASNase anaphylaxis, including anti-ASNase antibody responses, immune complex (IC) levels, ASNase-specific binding to leukocytes or basophils, and basophil activation.
RESULTS
Our results indicate that mice immunized to ASNase exhibited dynamic IgM, IgG, and IgE antibody responses. The severity of ASNase-induced anaphylaxis was found to be correlated with levels of IgG and IgE, but not IgM. Basophils from immunized mice were able to recognize and activate in response to ASNase ex vivo, and the extent of recognition and activation also correlated with the severity of anaphylaxis observed. Using a multivariable model that included all biomarkers significantly associated with anaphylaxis, independent predictors of ASNase-induced hypersensitivity reactions were found to be ASNase IC levels and ASNase-specific binding to leukocytes or basophils. Consistent with our multivariable analysis, we found that basophil depletion significantly protected mice from ASNase-induced hypersensitivity reactions, supporting that basophils are essential and can be used as a predictive marker of ASNase-induced anaphylaxis.
CONCLUSIONS
Our study demonstrates the need for using tools that can detect both IC- and IgE-mediated hypersensitivity reactions to mitigate the risk of ASNase-induced hypersensitivity reactions during treatment.
Topics: Animals; Asparaginase; Basophils; Mice; Drug Hypersensitivity; Anaphylaxis; Immunoglobulin E; Female; Disease Models, Animal; Biomarkers; Immunoglobulin G; Antineoplastic Agents
PubMed: 38686384
DOI: 10.3389/fimmu.2024.1392099 -
Journal of Microbiology and... May 2022L-asparaginase (E.C. 3.5.1.1) purified from bacterial cells is widely used in the food industry, as well as in the treatment of childhood acute lymphoblastic leukemia....
L-asparaginase (E.C. 3.5.1.1) purified from bacterial cells is widely used in the food industry, as well as in the treatment of childhood acute lymphoblastic leukemia. In the present study, the L-asparaginase gene was cloned into the pGEX-2T DNA plasmid, expressed in BL21 (DE3) pLysS, and purified to homogeneity using Glutathione Sepharose chromatography with 7.26 purification fold and 16.01% recovery. The purified enzyme exhibited a molecular weight of ~33.6 kDa with SDS-PAGE and showed maximal activity at 50°C and pH 8.0. It retained 95.1, 89.6%, and 70.2% initial activity after 60 min at 30°C, 40°C, and 50°C, respectively. The enzyme reserved its activity at 30°C and 37°C up to 24 h. The enzyme had optimum pH of 8 and reserved 50% activity up to 24 h. The recombinant enzyme showed the highest substrate specificity towards L-asparaginase substrate, while no detectable specificity was observed for L-glutamine, urea, and acrylamide at 10 mM concentration. THP-1, a human leukemia cell line, displayed significant morphological alterations after being treated with recombinant L-asparaginase and the IC of the purified enzyme was recorded as 0.8 IU. Furthermore, the purified recombinant L-asparaginase improved cytotoxicity in liver cancer HepG2 and breast cancer MCF-7 cell lines, with IC values of 1.53 and 18 IU, respectively.
Topics: Asparaginase; Burkholderia pseudomallei; Enzyme Stability; Escherichia coli; Humans; Recombinant Proteins; Substrate Specificity
PubMed: 35354764
DOI: 10.4014/jmb.2112.12050 -
Journal of Neuro-oncology Feb 2020Glioblastoma is an aggressive central nervous system tumor with a 5-year survival rate of < 10%. The standard therapy for glioblastoma is maximal safe resection,...
PURPOSE
Glioblastoma is an aggressive central nervous system tumor with a 5-year survival rate of < 10%. The standard therapy for glioblastoma is maximal safe resection, followed by radiation therapy and chemotherapy with temozolomide. New approaches to treatment of glioblastoma, such as targeting metabolism, have been studied. The object of this study is to evaluate whether asparagine could be a new target for treatment of glioblastoma.
METHODS
We investigated a potential treatment for glioblastoma that targets the amino acid metabolism. U251, U87, and SF767 glioblastoma cells were treated with L-asparaginase and/or 6-diazo-5-oxo-L-norleucine (DON). L-asparaginase hydrolyzes asparagine into aspartate and depletes asparagine. L-asparaginase has been used for the treatment of acute lymphoblastic leukemia. DON is a glutamine analog that inhibits several glutamine-utilizing enzymes, including asparagine synthetase.
RESULTS
Cell viability was measured after 72 h of treatment. MTS assay showed that L-asparaginase suppressed the proliferation of U251, U87, and SF767 cells in a dose-dependent manner. DON also inhibited the proliferation of these cell lines in a dose-dependent manner. Combined treatment with these drugs had a synergistic antiproliferative effect in these cell lines. Exogenous asparagine rescued the effect of inhibition of proliferation by L-asparaginase and DON. The expression of asparagine synthetase mRNA was increased in cells treated with a combination of L-asparaginase and DON. This combined treatment also induced greater apoptosis and autophagy than did single-drug treatment.
CONCLUSION
The results suggest that the combination of L-asparaginase and DON could be a new therapeutic option for patients with glioblastoma.
Topics: Antineoplastic Agents; Asparaginase; Brain Neoplasms; Cell Line, Tumor; Cell Proliferation; Cell Survival; Diazooxonorleucine; Glioblastoma; Humans
PubMed: 32020477
DOI: 10.1007/s11060-019-03351-4 -
World Journal of Microbiology &... Oct 2022This study aimed to select endophytic fungi to produce L-asparaginase and partially optimising the production of the enzyme using cacti as substrate. Seventeen...
This study aimed to select endophytic fungi to produce L-asparaginase and partially optimising the production of the enzyme using cacti as substrate. Seventeen endophytes were assessed for intracellular enzymatic potential in modified Czapek Dox's medium using L-proline as an inducer. The best producer was evaluated for intracellular and extracellular enzymatic activity in modified Czapek Dox's medium using flours of Opuntia ficus-indica and Nopalea cochenillifera as substrate. The biomass and L-asparaginase production profile was analysed and the best conditions for enzyme production were verified using factorial design. Penicillium decaturense URM 7966, Diaporthe ueckerae URM 8321, and Colletotrichum annellatum URM 8538 produced 0.76 U g, 0.87 U g, and 0.74 U g L-asparaginase, respectively. Diaporthe ueckerae URM 8321 produced only intracellular L-asparaginase, using flours of N. cochenillifera (0.72 U g) and O. ficus-indica (0.90 U g) and the last was selected for the next steps. The ideal time for biomass and L-asparaginase production was 120 h. The best conditions for enzyme production (1.67 U g) were initial pH 4.0, inoculum concentration 1% and cacti flour concentration 0.2%; where was observed an increase of 46.11% in compared to the initial production. Opuntia ficus-indica flour is indicated as an alternative low-cost substrate for the production of L-asparaginase by the endophytic fungus D. ueckerae URM 8321.
Topics: Asparaginase; Cactaceae; Fungi; Proline
PubMed: 36289148
DOI: 10.1007/s11274-022-03420-3 -
ESMO Open Sep 2020Insufficient exposure to asparaginase therapy is a barrier to optimal treatment and survival in childhood acute lymphoblastic leukaemia (ALL). Three important reasons... (Review)
Review
Insufficient exposure to asparaginase therapy is a barrier to optimal treatment and survival in childhood acute lymphoblastic leukaemia (ALL). Three important reasons for inactivity or discontinuation of asparaginase therapy are infusion related reactions (IRRs), pancreatitis and life-threatening central nervous system (CNS). For IRRs, real-time therapeutic drug monitoring (TDM) and premedication are important aspects to be considered. For pancreatitis and CNS thrombosis one key question is if patients should be re-exposed to asparaginase after their occurrence.An expert panel met during the Congress of the International Society for Paediatric Oncology in Lyon in October 2019 to discuss strategies for diminishing the impact of these three toxicities. The panel agreed that TDM is particularly useful for optimising asparaginase treatment and that when a tight pharmacological monitoring programme is established premedication could be implemented more broadly to minimise the risk of IRR. Re-exposure to asparaginase needs to be balanced against the anticipated risk of leukemic relapse. However, more prospective data are needed to give clear recommendations if to re-expose patients to asparaginase after the occurrence of severe pancreatitis and CNS thrombosis.
Topics: Antineoplastic Agents; Asparaginase; Child; Humans; Neoplasm Recurrence, Local; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Prospective Studies
PubMed: 32967920
DOI: 10.1136/esmoopen-2020-000977 -
Leukemia & Lymphoma Sep 2021is a widely explored genus due to its chemical diversity and associated biological properties; in addition, it represents an important source for cytotoxic compounds... (Review)
Review
is a widely explored genus due to its chemical diversity and associated biological properties; in addition, it represents an important source for cytotoxic compounds with good application perspectives. Based on these aspects, in this review, compounds that presented activity against human leukemia cell lines are being listed and discussed. For this, a careful bibliographic survey was carried out in the main electronic databases, i.e. Scopus, SciFinder, Web of Science and Pubmed. Between 1984 and 2020, thirty seven original papers were selected, when using the search terms and leukemia. The occurrence of l-asparaginase produced by some spp. was also highlighted since this enzyme is being employed for acute lymphoblastic leukemia and lymphosarcoma therapies. Therefore, this overview aims to demonstrate the potential of metabolites biosynthesized by fungi which can be applied in human leukemia therapies and opportunities for designing new lead compounds.
Topics: Antineoplastic Agents; Asparaginase; Humans; Leukemia; Penicillium
PubMed: 33733992
DOI: 10.1080/10428194.2021.1897804 -
Haematologica Oct 2023For several decades, asparaginase has been considered world-wide as an essential component of combination chemotherapy for the treatment of childhood acute lymphoblastic...
For several decades, asparaginase has been considered world-wide as an essential component of combination chemotherapy for the treatment of childhood acute lymphoblastic leukemia (ALL). Discovered over 60 years ago, two main unmanipulated asparaginase products originated from primary bacteria sources, namely Escherichia coli and Erwinia chrysanthemi, have been available for clinical use. A pegylated product of the Escherichia coli asparaginase was subsequently developed and is now the main product used by several international co-operative groups. The various asparaginase products all display the same mechanism of action (hydrolysis of circulating asparagine) and are associated with similar efficacy and toxicity patterns. However, their different pharmacokinetics, pharmacodynamics and immunological properties require distinctive modalities of application and monitoring. Erwinia chrysanthemi asparaginase was initially used as a first-line product, but subsequently became a preferred second-line product for children who experienced immunological reactions to the Escherichia coli asparaginase products. An asparaginase product displaying the same characteristics of the Erwinia chrysanthemi asparaginase has recently been produced by use of recombinant technology, thus securing a preparation available for use as an alternative, or as a back-up in case of shortages, for the non-recombinant product. The long journey of the Erwinia chrysanthemi asparaginase product as it has developed throughout the last several decades has made it possible for almost every child and adult with ALL to complete the asparaginase-based protocol treatment when an immunological reaction has occurred to any Escherichia coli asparaginase product.
Topics: Child; Adult; Humans; Asparaginase; Dickeya chrysanthemi; Drug Hypersensitivity; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Escherichia coli; Antineoplastic Agents
PubMed: 37470157
DOI: 10.3324/haematol.2022.282324