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A) 10% SDS-PAGE of isolated garlic alliinase; B) kinetic analysis of alliinase catalyzed reaction. The reaction mixture (0.1 mL) contained alliinase (5 μg/mL) and various concentrations of alliin (0 to 50 mM), the reaction temperature was 25˚C, Tricine-KOH (pH 8) buffer was used for the assays. https://doi.org/10.1371/journal.pone.0248878.g002
Source publication
Garlic is a well-known example of natural self-defence system consisting of an inactive substrate (alliin) and enzyme (alliinase) which, when combined, produce highly antimicrobial allicin. Increase of alliinase stability and its activity are of paramount importance in various applications relying on its use for in-situ synthesis of allicin or its...
Contexts in source publication
Context 1
... purified garlic alliinase, analyzed by 10% reducing SDS-PAGE showed a 51 kDa single band corresponding to alliinase monomer (Fig 2A), which is in agreement with previously published reports [33][34][35]. SDS-PAGE analysis revealed that alliinase obtained by PEG precipitation was of high purity without a significant presence of the accompanying proteins or degradation products. ...
Context 2
... lyophilized alliinase was found to have a protein content of 88 ± 4% per weight, accompanied by non-protein substances (polysaccharides, salts, PEG) and moisture. Kinetic parameters V max , K m and turnover number k cat for alliinase following MichaelisMenten kinetics with respect to the mixture of alliin diastereomers were calculated as 18.9 ± 0.3 mM�s -1 �mg -1 , 4.45 ± 0.36 mM and 193 s -1 , respectively (Fig 2B). The calculated values of kinetic parameters correspond to previously reported data [36]. ...
Citations
... aureus and anti-quorumsensing properties of MMOXC [38]. The stability of molecules that exhibit anti-microbial and anti-biofilm properties are evaluated for their stability in different conditions [39][40][41]. ...
... Also, each enzyme reaction and MMOXC complex was performed at pH 5.5 to 8.5, and at each pH, the absorbance was recorded. Similarly, for each enzyme and MMOXC complex increasing substrate concentration was followed by determining the activity at each substrate concentration [37,[39][40][41]46]. ...
... The continual rise in the drug-resistant bacteria globally has ushered scientists to develop new antibacterial drugs against which bacteria cannot develop defense machinery is the prime requirement to combat drug-resistant bacterial infections. The stability of such antibacterial compounds in extreme conditions such as elevated temperature, the increased time interval of incubation, pH, and elevated substrate concentration are the essential criteria for them to be considered as new drugs [38][39][40][41]46]. ...
The incidences of methicillin-resistant strains of Staphylococcus aureus (MRSA) and their survival inside the macrophages are the major attributes of the relapsed infections after antimicrobial therapy, and it is a global problem. In this context, we have previously demonstrated 4-methoxy-1-methyl-2-oxopyridine-3-carbamide (MMOXC), a Ricinine derivative exhibiting anti-S. aureus and anti-biofilm characteristics by competitively inhibiting uridine monophosphate kinase (UMPK), UDP-N-acetyl muramyl pentapeptide ligase (Mur-F), and peptidyl deformylase, (PDF). In the present study, the stability of this competitive inhibitor MMOXC was evaluated by showing its ability to remain bound to the active sites of UMPK, Mur-F, and PDF even after increasing the incubation time, temperature, pH, and substrate concentration. On growing MRSA in fewer concentrations of MMOXC, these strains could not attain resistance to MMOXC and at the same time distinct reductions in the expression of UMPK, Mur-F, and PDF genes were noted. In vitro, infective models were generated by infecting MRSA to RAW 264.7 and human monocyte-derived macrophage (hMDM) cell lines. In these infected cell lines, in spite of increased nitric oxide synthase (NOS), NADPH–P450 reductase, superoxide dismutase, catalase, and peroxidase activities, the MRSA survived. At 640 µM/ml, the concentration of MMOXC penetrated into these infected cells and obliterated MRSA. While treating uninfected macrophage cell lines with MMOXC, no appreciable effect was observed indicating that MMOXC is the most suitable drug for the treatment of infections caused by MRSA.
... However, the influence of additives on the properties of the enzyme is poorly understood because only very few systematic studies are available on this topic [8,9]. Hence, it is of pivotal importance to analyze the effect of various solvents, non-optimal pH, high ionic strength, inhibitors, and elevated temperature on enzyme structure and activity, all of which lead to enzyme denaturation and loss of activity [10,11]. Additives can either remarkably enhance or lower enzyme activity without causing any genetic or chemical modification [8,12]. ...
Interpreting the relationship between the activity and structure of β-galactosidase is necessary to perceive the impact of the enzyme’s conformation on its catalysis. The current study thoroughly explains the effects of additives such as ethylenediaminetetraacetic acid (EDTA), sodium dodecyl sulfate (SDS), dithiothreitol (DTT), and urea on β-galactosidase activity and structure. β-Galactosidase activity was determined at various ionic strengths and temperatures as a function of time. Structural studies evaluating changes in the secondary and tertiary structures of the enzyme in the presence of the additives were conducted using ultraviolet (UV)-visible and intrinsic fluorescence spectroscopy. The immobilized enzyme showed enhanced stability under different environmental conditions. Activity assays demonstrated concentration-dependent inactivation of β-galactosidase in the presence of SDS and urea, which suggests that hydrophobic and charged residues are present near the active site. In the presence of EDTA, loss in activity was noted, which confirms that β-galactosidase is a metalloenzyme. Enhancement in enzyme activity in the presence of DTT suggests the presence of a cysteine residue near the catalytic center. In UV-visible and intrinsic fluorescence spectroscopy studies, the native enzyme showed significant conformational transitions in the presence of DTT, SDS, and urea and very few changes in the presence of EDTA. However, the immobilized enzyme could resist significant structural changes. In conclusion, this study provides a detailed description of the association between the activity and conformational stability of β-galactosidase.
... As such, garlic with high alliinase activity is essential for the manufacture of garlic supplements. Alliinase constitutes up to 12% of garlic clove protein (Ellmore and Feldberg, 1994), and can be readily extracted from garlic, however it is not very stable due to the presence of free thiol groups which can become oxidised (Krest and Keusgen, 1999;Janská et al., 2021). Thus, for biotechnological extraction it is preferable to use fresh garlic bulbs. ...
Garlic (Allium sativum L.) has long been grown for its culinary and health-promoting qualities. The seasonal nature of garlic cropping requires that bulbs be stored for many months after harvest to ensure a year-round supply. During this time, quality is known to deteriorate, and efforts have been made to improve the longevity of stored bulbs. Cold temperatures within the stores prolong shelf life, but fine temperature control is needed to avoid freezing damage or cold induced stress. Here, quality traits (alliinase activity, firmness, and water content) are measured in response to a 96 h − 5 °C cold stress, to simulate the effect of non-isothermic temperature control in a − 1.5 °C warehouse. Volatile organic compounds (VOCs) are measured by thermal desorption gas chromatography time of flight mass spectrometry to identify markers of non-isothermic storage in garlic. 129 compounds were putatively identified and four (L-lactic acid, 2,6-dimethylhetpadecane, 4-methyldodecane, and methylcyclopentane) showed high predictive accuracy for cold stress. VOCs were also sampled directly from a cold storage facility and the whole profile discriminated between sampling time points. Five VOCS were highly predictive for storage time in the warehouse but were different to VOCs previously shown to discriminate between storage times in a laboratory setting. This indicates the need for realistic warehouse experiments to test quality markers.
... In rats, alliin treatment resulted in lower pathogenic antigen presence in the cecum, implying that alliin may suppress some opportunistic bacterial pathogens [28]. Finally, it is important to note that a natural antimicrobial agent formed from alliin and alliinase is allicin, which has been shown to damage several Gram-positive and Gram-negative bacterial strains like E. coli, P. putida and S. epidermidis [29]. ...
Background
Garlic has been considered to have medicinal properties in different cultures since ancient times. Vegetables of the allium genus are a natural source of sulfur-containing organic compounds. These compounds have recently been investigated for their positive health effects, such as antioxidant and anti-inflammatory, as well as cardioprotective and anticarcinogenic properties.
Objective
In this review we focus on one of the main components of garlic; alliin or S-allyl-L-cysteine sulfoxide, which is a non-protein amino acid that exhibits a broad spectrum of beneficial effects on physiology, both at the cellular and whole organism levels.
Methods
Methods: We conducted a systematic literature search of the MEDLINE (PubMed) database. Search terms used for alliin were: “S-allylcysteine sulfoxide” OR “L-alliin” OR “ACSO”, all of them combined into a separated search term individually as follows: AND “antioxidant”; AND “cardioprotective”; AND “anti-inflammatory”; AND “antimicrobial”; AND “disease”, and; AND “neuroprotective”.
Results
Here, we review and integrate the existing experimental evidence on the effects of alliin, mainly on its antioxidant and anti-inflammatory effects, as well as its cardioprotective action, and its role as an adjuvant for the treatment of different diseases, such as: infectious diseases, inflammatory diseases, metabolic diseases and cancer.
Conclusion
Finally, we propose alliin as a possible neuroprotective agent, through the combination of its antioxidant and anti-inflammatory effects, and its ability to reduce markers of metabolic inflammation in obesity.
... The disulfide bond, which is the main resource of the antibacterial properties in ALL, is particularly prone to rupture under alkaline conditions (Wills, 1956). One popular approach to solve this problem is to encapsulate allicin with proper encapsulation (Lawson and Hughes, 1992;Janská et al., 2021). For example, Malgorzata et al. designed oil-core nanocapsules based on a derivative of hyaluronic acid to protect garlic oil active components, resulting in preventing sulfur oxidation and maintaining antibacterial activities (Janik-Hazuka et al., 2021). ...
Frontiers e-book, comprising all the articles featured in the Research Topic "Advances in Materials Toward Anti-Corrosion and Anti-Biofoulings". Alternatively, you can also access the e-book via the Research Topic homepage (https://www.frontiersin.org/research-topics/24136/advances-in-materials-toward-anti-corrosion-and-anti-biofoulings), where all articles are also individually available.
... The disulfide bond, which is the main resource of the antibacterial properties in ALL, is particularly prone to rupture under alkaline conditions (Wills, 1956). One popular approach to solve this problem is to encapsulate allicin with proper encapsulation (Lawson and Hughes, 1992;Janská et al., 2021). For example, Malgorzata et al. designed oil-core nanocapsules based on a derivative of hyaluronic acid to protect garlic oil active components, resulting in preventing sulfur oxidation and maintaining antibacterial activities (Janik-Hazuka et al., 2021). ...
In this work, we report the design of pH-controlled releasing behaviors of polydopamine/tannic acid-allicin@chitosan (PDA/TA-ALL@CS) multilayer coatings to realize antibacterial and antifouling effects. The pH-responsive ALL@CS capsules were prepared using the microemulsion method with about 262–452 nm diameter. The bacteriostasis of ALL@CS microcapsules against E. coli , S. aureus , and P. aeruginosa all exceeded 94% as evaluated using the colony counting method. Because of the protonation in acid environments and deprotonation in alkaline environments for the amino groups of CS, ALL as biocides can be released from the nanocapsules and exert outstanding antibacterial properties. Confirmed by the plate colony counts, the ALL@CS capsules possessed an outstanding antibacterial effect for E. coli in acid solutions but were less effective in alkaline solutions. The PDA/TA-ALL@CS-7 coatings showed durable pH-responsive antibacterial activities with an efficiency of ∼87% after immersion in pH 8 solutions for seven days. The PDA/TA-ALL@CS coating with controlled release performance and antibacterial properties may provide a new solution for developing antifouling coating applications in the marine environment.
... Alliinase was extracted from fresh garlic according to previously published protocols with minor modifications [29,30]. The whole isolation procedure was carried out at 4°C. ...
... In this way, the native conformation of the protein is maintained even in a solid state [48]. Moreover, the negative effect of moisture, which is 4-times higher in chitosan compared to the maltodextrin particles, on protein stability is well documented [30,49]. Thereby, the protective effect of maltodextrin can be further magnified, lowering the moisture content in the final product. ...
Allicin, an organic compound produced via the transformation of biologically inactive alliin by alliinase, an enzyme found in garlic, combines a strong antibacterial effect with suppressed development of bacterial resistance. However, because of the high reactivity and volatility, controlled in-situ production of allicin that mimics the natural synthesis is essential to achieve desired therapeutic effects. In this work, the spray-drying technique was employed for encapsulation of alliinase into polymer micro-carriers with an emphasis on the effect of process parameters, i.e., drying temperature, nozzle type, choice of the carrier materials, on the activity of encapsulated alliinase in soluble maltodextrin and swellable chitosan microparticles. The results show that maltodextrin is a suitable carrier for the effective protection of alliinase against thermal stress. On the other hand, we can control allicin production and release from chitosan carriers with immobilized alliinase by variation of the cross-linker amount. Antibacterial activity of in-situ formed allicin vapors was confirmed against Escherichia coli bacterial strain using customized sample holders preventing physical contact of powder sample with the inoculated agar plate.
Escherichia coli (E. coli) strains cause the majority of urinary tract infections (UTIs) and are resistant to various antibiotics. Therefore, it is imperative to explore novel host-target therapies. As a famous food and condiment, garlic (Allium sativum L.) is widely used in medicine, but its exact key targets in UTIs remain elusive. To identify the major active ingredient of garlic and its molecular target against UTIs, a network pharmacology analysis was carried out, and allicin was revealed to be a key active component in garlic acting on UTIs. By molecular docking, allicin showed a good binding affinity to mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1). The possible regulatory mechanisms of allicin against UTIs were based on the modules of immune and inflammatory responses mainly through AKT/NF-κB signaling. Next, an E. coli-stimulated human uroepithelial cell (HUC) model was established to confirm the anti-infective effect of allicin. The results showed that allicin could significantly inhibit the upregulation of MALT1, the AKT/NF-κB pathway, and cytokines (IL-6 and IL-1β). HUCs pretreated with the PI3K inhibitor or transfected with MALT-siRNA also partly suppressed the activation of the AKT/NF-κB pathway and cytokines. Furthermore, by establishing the PCA algorithm to evaluate the therapeutic score, allicin was proved to achieve the optimal therapeutic effects compared with the PI3K inhibitor and siRNA-MALT1. Moreover, in rats with an E. coli-induced UTI model, allicin significantly alleviated the infection and up-regulation of MALT1 expression in the bladders, a marked increase in the bacterial load of urine, and deviations in serum biochemical parameters. In conclusion, allicin exerts anti-infective effects in UTIs mainly via the MALT1/NF-κB axis or AKT/NF-κB pathway, which provides a theoretical basis for understanding the function of allicin against UTIs and facilitates its clinical application.
Recently, the consumption of hummus has become popular in the United States, European countries, and Canada, and unfortunately, several foodborne outbreaks and recalls have been reported due to its contamination with Listeria monocytogenes and Salmonella enterica. The current study aimed to investigate the inhibitory activity of 0.5, 1.0, and 1.5% citric acid (CA) and 1.0, 2.0 and 3.0% garlic extract (GE) toward S. enterica and L. monocytogenes in hummus stored at 4, 10 and 24 °C. L. monocytogenes grew well in untreated (control) hummus samples at all tested temperatures, whereas S. enterica grew only at 10 and 24 °C. CA at 0.5 to 1.5% reduced L. monocytogenes numbers by 3.0–3.3 log CFU/g at 4 °C, 1.7–3.9 log CFU/g at 10 °C, and 0.9–1.4 log CFU/g at 24 °C. Numbers of S. enterica were reduced by 0.6–1.7, 4.1–4.9 and <1.5 log CFU/g, at 4, 10 and 24 °C, respectively, compared to the control during 10 d storage. GE at 1.0–3.0% also reduced numbers of L. monocytogenes at 10 d by 0.7–3.0, and 1.3–3.6 log CFU/g at 4 and 10 °C, respectively, and numbers of S. enterica by 0.7–1.2, 1.8–2.6 and 0.5–1.6 log CFU/g, at 4, 10 and 24 °C, respectively, compared to the control. Chromatographic analysis of GE revealed the presence four organosulfur compounds including diallyl disulfide, diallyl trisulfide, 2-vinyl-(4H)-1,3-dithiin and 3-vinyl-(4H)-1,2-dithiin where the latter was the predominant compound with a level of 22.9 mg/g which mainly contributed to the inhibitory effect of GE. CA and GE are adequate natural antimicrobials in hummus to reduce L. monocytogenes and S. enterica numbers and enhance product safety.
