Tea crude extracts effectively inactivate severe acute respiratory syndrome coronavirus 2.

It is well known that black and green tea extracts, particularly polyphenols, have antimicrobial activity against various pathogenic microbes including viruses. However, there is limited data on the antiviral activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which emerged rapidly in China in late 2019 and which has been responsible for coronavirus disease 2019 (COVID-19) pandemic globally. In this study, 20 compounds and three extracts were obtained from black and green tea and found that three tea extracts showed significant antiviral activity against SARS-CoV-2, whereby the viral titre decreased about 5 logs TCID50 per ml by 1·375 mg ml-1 black tea extract and two-fold diluted tea bag infusion obtained from black tea when incubated at 25°C for 10 s. However, when concentrations of black and green tea extracts were equally adjusted to 344 µg ml-1 , green tea extracts showed more antiviral activity against SARS-CoV-2. This simple and highly respected beverage may be a cheap and widely acceptable means to reduce SARS-CoV-2 viral burden in the mouth and upper gastrointestinal and respiratory tracts in developed as well as developing countries.

Chlorine dioxide is a more potent antiviral agent against SARS-CoV-2 than sodium hypochlorite.

BACKGROUND: A new coronavirus (SARS-CoV-2) abruptly emerged in Wuhan, China, in 2019 and rapidly spread globally to cause the COVID-19 pandemic. AIM: To examine the anti-SARS-CoV-2 activity of the potent disinfectant Cleverin, the major disinfecting component of which is chlorine dioxide (ClO2); and to compare the results with that of sodium hypochlorite in the presence or absence of 0.5% or 1.0% foetal bovine serum (FBS). METHODS: Concentrated SARS-CoV-2 viruses were treated with various concentrations of ClO2 and sodium hypochlorite and 50% tissue culture infective dose was calcurated to evaluate the antiviral activity of each chemical. FINDINGS: When SARS-CoV-2 viruses were treated with 0.8 ppm ClO2 or sodium hypochlorite, viral titre was decreased only by 1 log10 TCID50/mL in 3 min. However, the viral titre was decreased by more than 4 log10 TCID50/mL when treated with 80 ppm of each chemical for 10 s regardless of presence or absence of FBS. It should be emphasized that treatment with 24 ppm of ClO2 inactivated more than 99.99% SARS-CoV-2 within 10 s or 99.99% SARS-CoV-2 in 1 min in the presence of 0.5% or 1.0% FBS, respectively. By contrast, 24 ppm of sodium hypochlorite inactivated only 99% or 90% SARS-CoV-2 in 3 min under similar conditions. Notably, except for ClO2, the other components of Cleverin such as sodium chlorite, decaglycerol monolaurate, and silicone showed no significant antiviral activity. CONCLUSION: Altogether, the results strongly suggest that although ClO2 and sodium hypochlorite are strong antiviral agents in absence of organic matter but in presence of organic matter, ClO2 is a more potent antiviral agent against SARS-CoV-2 than sodium hypochlorite.

Analysis of the effect of accumulation of amino acid replacements on activity of 3-isopropylmalate dehydrogenase from Thermus thermophilus.

A newly selected cold-adapted mutant 3-isopropylmalate dehydrogenase (IPMDH) from a random mutant library was a double mutant containing the mutations I11V and S92F that were found in cold-adapted mutant IPMDHs previously isolated. To elucidate the effect of each mutation on enzymatic activity, I11V and six multiple mutant IPMDHs were constructed and analyzed. All of the multiple mutant IPMDHs were found to be improved in catalytic activity at moderate temperatures by increasing the k(cat) with a simultaneous increase of K(m) for the coenzyme NAD(+). k(cat) was improved by a decrease in the activation enthalpy, DeltaH( not equal). The multiple mutants did not show large reduction in thermal stability, and one of them showed enhanced thermal stability. Mutation from I11 to V was revealed to have a stabilizing effect. Mutants showed increased thermal stability when the mutation I11V was combined. This indicates that it is possible to construct mutants with enhanced thermal stability by combining stabilizing mutation. No additivity was observed for the thermodynamic properties of catalytic reaction in the multiple mutant IPMDHs, implying that the structural changes induced by the mutations were interacting with each other. This indicates that careful and detailed tuning is required for enhancing activity in contrast to thermal stability.

Adaptation of a thermophilic enzyme, 3-isopropylmalate dehydrogenase, to low temperatures.

Random mutagenesis coupled with screening of the active enzyme at a low temperature was applied to isolate cold-adapted mutants of a thermophilic enzyme. Four mutant enzymes with enhanced specific activities (up to 4.1-fold at 40 degrees C) at a moderate temperature were isolated from randomly mutated Thermus thermophilus 3-isopropylmalate dehydrogenase. Kinetic analysis revealed two types of cold-adapted mutants, i.e. k(cat)-improved and K(m)-improved types. The k(cat)-improved mutants showed less temperature-dependent catalytic properties, resulting in improvement of k(cat) (up to 7.5-fold at 40 degrees C) at lower temperatures with increased K(m) values mainly for NAD. The K(m)-improved enzyme showed higher affinities toward the substrate and the coenzyme without significant change in k(cat) at the temperatures investigated (30-70 degrees C). In k(cat)-improved mutants, replacement of a residue was found near the binding pocket for the adenine portion of NAD. Two of the mutants retained thermal stability indistinguishable from the wild-type enzyme. Extreme thermal stability of the thermophilic enzyme is not necessarily decreased to improve the catalytic function at lower temperatures. The present strategy provides a powerful tool for obtaining active mutant enzymes at lower temperatures. The results also indicate that it is possible to obtain cold-adapted mutant enzymes with high thermal stability.

Cold adaptation of the thermophilic enzyme 3-isopropylmalate dehydrogenase.

We have performed random mutagenesis coupled with selection to isolate mutant enzymes with high catalytic activities at low temperature from thermophilic 3-isopropylmalate dehydrogenase (IPMDH) originally isolated from Thermus thermophilus. Five cold-adapted mutant IPMDHs with single-amino-acid substitutions were obtained and analyzed. Kinetic analysis revealed that there are two types of cold-adapted mutant IPMDH: k(cat)-improved (improved in k(cat)) and K(m)-improved (improved in k(cat)/K(m)) types. To determine the mechanisms of cold adaptation of these mutants, thermodynamic parameters were estimated and compared with those of the Escherichia coli wild-type IPMDH. The Delta G(m) values for Michaelis intermediate formation of the k(cat)-improved-type enzymes were larger than that of the T. thermophilus wild-type IPMDH and similar to that of the E. coli wild-type IPMDH. The Delta G(m) values of K(m)-improved-type enzymes were smaller than that of the T. thermophilus wild-type IPMDH. Fitting of NAD(+) binding was improved in the K(m)-improved-type enzymes. The two types of cold-adapted mutants employed one of the two strategies of E. coli wild-type IPMDH: relative destabilization of the Michaelis complex in k(cat)-improved-type, and destabilization of the rate-limiting step in K(m)-improved type mutants. Some cold-adapted mutant IPMDHs retained thermostability similar to that of the T. thermophilus wild-type IPMDH.