Phycobilins as Potent Food Bioactive Broad-Spectrum Inhibitors Against Proteases of SARS-CoV-2 and Other Coronaviruses: A Preliminary Study

In the 21st century, we have witnessed three coronavirus outbreaks: SARS in 2003, MERS in 2012, and the ongoing pandemic coronavirus disease 2019 (COVID-19). The search for efficient vaccines and development and repurposing of therapeutic drugs are the major approaches in the COVID-19 pandemic research area. There are concerns about the evolution of mutant strains (e.g., VUI – 202012/01, a mutant coronavirus in the United Kingdom), which can potentially reduce the impact of the current vaccine and therapeutic drug development trials. One promising approach to counter the mutant strains is the “development of effective broad-spectrum antiviral drugs” against coronaviruses. This study scientifically investigates potent food bioactive broad-spectrum antiviral compounds by targeting main protease (Mpro) and papain-like protease (PLpro) proteases of coronaviruses (CoVs) using in silico and in vitro approaches. The results reveal that phycocyanobilin (PCB) shows potential inhibitor activity against both proteases. PCB had the best binding affinity to Mpro and PLpro with IC50 values of 71 and 62 μm, respectively. Also, in silico studies with Mpro and PLpro enzymes of other human and animal CoVs indicate broad-spectrum inhibitor activity of the PCB. As with PCB, other phycobilins, such as phycourobilin (PUB), phycoerythrobilin (PEB), and phycoviolobilin (PVB) show similar binding affinity to SARS-CoV-2 Mpro and PLpro

Genomic Modeling as an Approach to Identify Surrogates for Use in Experimental Validation of SARS-CoV-2 and HuNoV Inactivation by UV-C Treatment

Severe Acute Respiratory Syndrome coronavirus-2 (SARS-CoV-2) is responsible for the COVID-19 pandemic that continues to pose significant public health concerns. While research to deliver vaccines and antivirals are being pursued, various effective technologies to control its environmental spread are also being targeted. Ultraviolet light (UV-C) technologies are effective against a broad spectrum of microorganisms when used even on large surface areas. In this study, we developed a pyrimidine dinucleotide frequency based genomic model to predict the sensitivity of select enveloped and non-enveloped viruses to UV-C treatments in order to identify potential SARS-CoV-2 and human norovirus surrogates. The results revealed that this model was best fitted using linear regression with r2 = 0.90. The predicted UV-C sensitivity (D90 – dose for 90% inactivation) for SARS-CoV-2 and MERS-CoV was found to be 21.5 and 28 J/m2, respectively (with an estimated 18 J/m2 obtained from published experimental data for SARS-CoV-1), suggesting that coronaviruses are highly sensitive to UV-C light compared to other ssRNA viruses used in this modeling study. Murine hepatitis virus (MHV) A59 strain with a D90 of 21 J/m2 close to that of SARS-CoV-2 was identified as a suitable surrogate to validate SARS-CoV-2 inactivation by UV-C treatment. Furthermore, the non-enveloped human noroviruses (HuNoVs), had predicted D90 values of 69.1, 89, and 77.6 J/m2 for genogroups GI, GII, and GIV, respectively. Murine norovirus (MNV-1) of GV with a D90 = 100 J/m2 was identified as a potential conservative surrogate for UV-C inactivation of these HuNoVs. This study provides useful insights for the identification of potential non-pathogenic (to humans) surrogates to understand inactivation kinetics and their use in experimental validation of UV-C disinfection systems. This approach can be used to narrow the number of surrogates used in testing UV-C inactivation of other human and animal ssRNA viral pathogens for experimental validation that can save cost, labor and time

UV-C inactivation of microorganisms in droplets on food contact surfaces using UV-C light-emitting diode devices

The main objective of this study was to investigate the effectiveness of ultraviolet light (UV-C) emitting diodes for the decontamination of stainless steel food contact surfaces. Listeria monocytogenes (ATCC 19115), Escherichia coli (ATCC 25922), and Salmonella enterica serovar Typhimurium (ATCC 700720) were chosen as challenge microorganisms. Target microorganisms were subjected to UV-C dosages of 0, 2, 4, 6, and 8 mJ cm−2 at an average fluence of 0.163 mW/cm2 using a near-collimated beam operating at 279 nm wavelength. Escherichia coli showed lower sensitivity to UV-C light compared to Salmonella Typhimurium and followed first-order kinetics. Escherichia coli and Salmonella Typhimurium were reduced by more than 3-log10 cycles at the maximum UV dosage of 12 mJ cm−2 . In contrast, Listeria monocytogenes followed the Weibull model with an apparent shoulder in the initial doses. A maximum reduction of 4.4-log10 was achieved at the highest exposure level. This study showed that UV-C LED devices represent an excellent alternative for the inactivation of foodborne microorganisms in droplets. Results clearly demonstrate that UV-C LED devices can serve as an additional sanitation method to routine cleaning practices, which are commonly utilized in the food industry

Atmospheric Cold Plasma Inactivation of Salmonella and Escherichia coli on the Surface of Golden Delicious Apples

The contamination of fruits with human pathogens is a reoccurring concern in the fresh produce industry. Atmospheric cold plasma (ACP) is a potential alternate to customary approaches for non-thermal decontamination of foods. In this study, the efficacy of a dielectric barrier discharge ACP system against Salmonella (Salmonella Typhimurium, ATCC 13311; Salmonella Choleraesuis, ATCC 10708) and Escherichia coli (ATCC 25922, ATCC 11775) was explored. For each bacteria, a two-strain mixture at 8 log10 CFU/ml was spot inoculated on the surface of Golden Delicious apples, air dried, and exposed to ACP at a fixed distance of 35 mm, input power of 200 W for 30, 60, 120, 180, and 240 s. Bacterial inactivation was achieved in all treatment times with highest reduction of 5.3 log10 CFU/cm2 for Salmonella and 5.5 log10 CFU/cm2 for E. coli. Our results showed that reductions were interrelated to exposure time and ranged from 1.3 to 5.3 and 0.6 to 5.5 log10 CFU/cm2 for Salmonella and E. coli, respectively. Salmonella and E. coli significantly decreased (\u3e5.0 log) at 180 and 240 s as compared to 30, 60, and 120 s exposure. Microbial inactivation data was modeled by using Weibull distribution. These findings demonstrate the potential of ACP as a postharvest technology to effectively reduce pathogens on apples, with reference to Salmonella and E. coli

Phycobilins as Potent Food Bioactive Broad-Spectrum Inhibitors Against Proteases of SARS-CoV-2 and Other Coronaviruses: A Preliminary Study

In the 21st century, we have witnessed three coronavirus outbreaks: SARS in 2003, MERS in 2012, and the ongoing pandemic coronavirus disease 2019 (COVID-19). The search for efficient vaccines and development and repurposing of therapeutic drugs are the major approaches in the COVID-19 pandemic research area. There are concerns about the evolution of mutant strains (e.g., VUI – 202012/01, a mutant coronavirus in the United Kingdom), which can potentially reduce the impact of the current vaccine and therapeutic drug development trials. One promising approach to counter the mutant strains is the “development of effective broad-spectrum antiviral drugs” against coronaviruses. This study scientifically investigates potent food bioactive broad-spectrum antiviral compounds by targeting main protease (Mpro) and papain-like protease (PLpro) proteases of coronaviruses (CoVs) using in silico and in vitro approaches. The results reveal that phycocyanobilin (PCB) shows potential inhibitor activity against both proteases. PCB had the best binding affinity to Mpro and PLpro with IC50 values of 71 and 62 μm, respectively. Also, in silico studies with Mpro and PLpro enzymes of other human and animal CoVs indicate broad-spectrum inhibitor activity of the PCB. As with PCB, other phycobilins, such as phycourobilin (PUB), phycoerythrobilin (PEB), and phycoviolobilin (PVB) show similar binding affinity to SARS-CoV-2 Mpro and PLpro

Performance of a UV-A LED system for degradation of aflatoxins B1 and M1 in pure water: kinetics and cytotoxicity study

The efficacy of a UV-A light emitting diode system (LED) to reduce the concentrations of aflatoxin B1, aflatoxin M1 (AFB1, AFM1) in pure water was studied. This work investigates and reveals the kinetics and main mechanism(s) responsible for the destruction of aflatoxins in pure water and assesses the cytotoxicity in liver hepatocellular cells. Irradiation experiments were conducted using an LED system operating at 365 nm (monochromatic wave-length). Known concentrations of aflatoxins were spiked in water and irradiated at UV-A doses ranging from 0 to 1,200 mJ/cm2. The concentration of AFB1 and AFM1 was determined by HPLC with fluorescence detection. LC–MS/MS product ion scans were used to identify and semi-quantify degraded products of AFB1 and AFM1. It was observed that UV-A irradiation significantly reduced aflatoxins in pure water. In comparison to control, at dose of 1,200 mJ/cm2 UV-A irradiation reduced AFB1 and AFM1 concentrations by 70 ± 0.27 and 84 ± 1.95%, respectively. We hypothesize that the formation of reactive species initiated by UV-A light may have caused photolysis of AFB1 and AFM1 molecules in water. In cell culture studies, our results demonstrated that the increase of UV-A dosage decreased the aflatoxins-induced cytotoxicity in HepG2 cells, and no significant aflatoxin-induced cytotoxicity was observed at UV-A dose of 1,200 mJ/cm2. Further results from this study will be used to compare aflatoxins detoxification kinetics and mechanisms involved in liquid foods such as milk and vegetable oils

Efficacy of ultraviolet (UV-C) light in reducing foodborne pathogens and model viruses in skim milk

The efficacy of low wavelength ultraviolet light (UV-C) as a disinfection process for a scattering fluid such as skim milk was investigated in this study. UV-C inactivation kinetics of two surrogate viruses (bacteriophages MS2 and T1UV) and three bacteria Escherichia coli ATCC 25922, S. Typhimurium ATCC 13311, Listeria monocytogenes ATCC 19115 in buffer and skim milk were investigated. UV-C irradiation was applied to stirred samples, using a collimated beam operating at 253.7 nm wavelength. A series of known UV-C doses (0–40 mJ·cm−2) were delivered to the samples except MS2 where higher doses (0–150 mJ·cm−2) were delivered. Biodosimetry, utilizing D values of viruses inactivated in buffer, was carried out to verify and calculate reduction equivalent dose. At the highest dose of 40 mJ·cm−2, the three pathogenic organisms were inactivated by more than 5 log10 (p \u3c .05). Results provide evidence that UV-C irradiation effectively inactivated bacteriophage and pathogenic microbes in skim milk. The inactivation kinetics of microorganisms was well described by log linear and exponential models with a low root mean squared error and high coefficient of determination (r2 \u3e 0.96). Models were validated and parameterized for predicting log reduction as a function of UV-C irradiation dose (p \u3c .05). This study clearly demonstrated that high levels of inactivation of pathogens can be achieved in skim milk, and suggests significant potential for UV-C treatment of treating fluids that exhibit significant scattering. Practical application This research paper provides scientific evidence of the potential use of UV technology in inactivating pathogenic bacteria and model viruses in skim milk. UV-C doses were validated and verified using biodosimetry. UV-C irradiation is an attractive food preservation technology and offers opportunities for dairy and food processing industries to meet the growing demand from consumers for safer food products. This study clearly shows the potential for using UV-C treatment for treating highly scattering fluid such as skim milk. Results from this work will be used to further develop continuous flow-through UV-C systems based on dean or turbulent flow patterns

UV-C treatment on the safety of skim milk: Effect on microbial inactivation and cytotoxicity evaluation

The efficacy of UV-C irradiation as a nonthermal processing method for skim milk (SM) was investigated. SM inoculated with two surrogate viruses (MS2 and T1UV), and three bacteria (Escherichia coli ATCC 25922, Salmonella enterica serovar Typhimurium ATCC 13311, and Listeria monocytogenes ATCC 19115) was treated with Deanflow UV-C irradiation, a spiral reactor with the fluid pumped around a central low-pressure mercury UV lamp (40 W) emitting at 254 nm wave-length. A series of known UV doses (0–168.33 mJ·cm2) were delivered to the samples. The microbial loads of MS2, T1UV, E. coli, Salmonella, and Listeria were reduced by more than 5 log10. The inactivation kinetics of all microorganisms were best described by log linear models with a low RMSE and higher coefficient of determination (R2 \u3e 0.95). This study demonstrated that high levels of inactivation of pathogenic particles can be achieved in SM. Practical applications This scientific study provides evidence based data on the advantages of UV-C light in achieving microbial reduction in skim milk. The irradiated skim milk did not show any toxicity on mice liver and intestinal cells. UV-C irradiation is an efficient food preservation technology and offers opportunities for dairy and food processing industries to meet the growing demand from consumers for safer foods. This exploration would provide methodological evidence for commercialization of UV-C processing of milk and dairy based beverages

Optimization of Ultrasound Assisted Extraction of Antioxidant Compounds from Marjoram (Origanum majorana L.) Using Response Surface Methodology

The present study optimized the ultrasound assisted extraction (UAE) conditions to maximize the antioxidant activity [Ferric ion Reducing Antioxidant Power (FRAP)], total phenol content (TP) and content of individual polyphenols of extracts from marjoram. Optimal conditions with regard to amplitude of sonication (24.4–61.0 μm) and extraction temperature (15–35 °C) and extraction time (5–15 min) were identified using response surface methodology (RSM). The results showed that the combined treatment conditions of 61 μm, 35 °C and 15 min were optimal for maximizing TP, FRAP, rosmarinic acid, luteolin-7-O-glucoside, apigenin-7-O-glucoside, caffeic acid, carnosic acid and carnosol values of the extracts. The predicted values from the developed quadratic polynomial equation were in close agreement with the actual experimental values with low average mean deviations (E%) ranging from 0.45% to 1.55%. The extraction yields of the optimal UAE were significantly (p \u3c 0.05) higher than solid/liquid extracts. Predicted models were highly significant (p \u3c 0.05) for all the parameters studied with high regression coefficients (R2) ranging from 0.58 to 0.98

Microbial inactivation and cytotoxicity evaluation of UV irradiated coconut water in a novel continuous flow spiral reactor

A continuous-flow UV reactor operating at 254 nm wave-length was used to investigate inactivation of microorganisms including bacteriophage in coconut water, a highly opaque liquid food. UV-C inactivation kinetics of two surrogate viruses (MS2, T1UV) and three bacteria (E. coli ATCC 25922, Salmonella Typhimurium ATCC 13311, Listeria monocytogenes ATCC 19115) in buffer and coconut water were investigated (D10 values ranging from 2.82 to 4.54 mJ·cm− 2). A series of known UV-C doses were delivered to the samples. Inactivation levels of all organisms were linearly proportional to UV-C dose (r2 \u3e 0.97). At the highest dose of 30 mJ·cm− 2, the three pathogenic organisms were inactivated by \u3e 5 log10 (p \u3c 0.05). Results clearly demonstrated that UV-C irradiation effectively inactivated bacteriophage and pathogenic microbes in coconut water. The inactivation kinetics of microorganisms were best described by log linear model with a low root mean square error (RMSE) and high coefficient of determination (r2 \u3e 0.97). Models for predicting log reduction as a function of UV-C irradiation dose were found to be significant (p \u3c 0.05) with low RMSE and high r2. The irradiated coconut water showed no cytotoxic effects on normal human intestinal cells and normal mouse liver cells. Overall, these results indicated that UV-C treatment did not generate cytotoxic compounds in the coconut water. This study clearly demonstrated that high levels of inactivation of pathogens can be achieved in coconut water, and suggested potential method for UV-C treatment of other liquid foods. Industrial relevance This research paper provides scientific evidence of the potential benefits of UV-C irradiation in inactivating bacterial and viral surrogates at commercially relevant doses of 0–120 mJ·cm− 2. The irradiated coconut water showed no cytotoxic effects on normal intestinal and healthy mice liver cells. UV-C irradiation is an attractive food preservation technology and offers opportunities for horticultural and food processing industries to meet the growing demand from consumers for healthier and safe food products. This study would provide technical support for commercialization of UV-C treatment of beverages