Progress 08/15/24 to 08/14/25

Outputs
Target Audience:Scientists, professionals, and technical personals in the nanotech for food preservation and safety research community, and also decision makers, engineers, and other technical personnel in the relevant industry searching for and/or working on new or more effective food preservation and safety technologies. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?We have continued our major effort on providing hands-on laboratory research trainings for students and all team members in a highly interdisciplinary and cross-institutional project environment. The students at graduate and undergraduate levels have benefited significantly from their participation in the project. One graduate student so inspired by her participation in this project decided to compete for the USDA/NIFA pre-doctoral fellowship with ideas related to but beyond those behind this project for expanded food safety applications. All participating graduate students and also undergraduate student researchers have been excited about their performing the project tasks and getting good results for their theses and related manuscripts for publication. The recent graduates from our programs who were trained well in and benefited majorly from this project and are now in R&D jobs or in other graduate programs have become role models for the current students to be well prepared for pursuing similar opportunities in relevant technical fields. How have the results been disseminated to communities of interest?The research activities in this project have become a critical part of our overall effort in the development of carbon dots and their derived nanotechnology. We have been actively disseminating the results from our continuing effort. The peer-reviewed journal publications acknowledging USDA support via this project are as follows. Collins, J.; Yang, L.; Dong, X.; Sun, Y.-P. "Antimicrobial Properties of Carbon "Quantum" Dots for Food Safety Applications." J. Nanoparticle Res., 2025, 27(2), article #35. (DOI: 10.1007/s11051-025-06239-9) Sun, Y.-P.; Yang, L. "Carbon Dots from Preexisting Nanoparticles versus Carbonization - Similarity and Difference in Sample Structure-Morphology with Property and Mechanistic Consequences." Small Structures 2025, 6(3), 2400455. (DOI: 10.1002/sstr.202400455) Singh, B.; Adcock, A. F.; Dumra, S.; Collins, J.; Yang, L.; Bunker, C. E.; Qian, H.; Meziani, M. J.; Sun, Y.-P. "Microwave-Assisted Carbonization Processing for Carbon Dots-Like Nanomaterials of Antimicrobial Properties." Micro 2025, 5, 14. (DOI: 10.3390/micro5010014) Sheriff, K.; Liang, W.; Yerra, S.; Sulejmanovic, D.; Nelson, C.; Lupini, A.; Hwu, S.-J.; Singh, B.; Sun, Y.-P. "Carbon Dots from Surface-Capping/Passivation of Small Carbon Nanoparticles with Nanoscale Titanium Dioxide." Chem. Phys. Lett. 2025, 864, 141913. (DOI: 10.1016/j.cplett.2025.141913) Rodriguez, C.; Adcock, A. F.; Dong, X.; Yerra, S.; Sun, Y.-P.; Yang, L. "Antimicrobial Activity of Carbon Dots in Combination with Different Co-Agents and Mechanistic Implications." J. Nanoparticle Res. 2025, revised manuscript waiting for the editor's decision. What do you plan to do during the next reporting period to accomplish the goals?We have had two successful years for our project, which is clearly on the right track and on schedule, so we will continue and expand our progress in the performance of the project tasks. In the next project year, we will complete most of the 1st and 2nd tasks and some of the 3rd task in Aim 1, continue the tasks in Aim 2, and ramp up our effort on the tasks in Aim 3. We will disseminate as much as possible the results from our project to the nanotech for food preservation and safety research community through peer-reviewed publications and conference presentations. Of equal importance is our continuing focus on providing our graduate and undergraduate students with excellent trainings in the related research areas.

Impacts
What was accomplished under these goals? We have built upon the significant progress made in year-1 to continue the project tasks for Aims 1 & Aim 2 and also to get ready for Aim 3. 1. More Significant Progress in Food-Safe Carbon Dots Platform. Classical carbon dots (CDots) are small carbon nanoparticles (CNPs) with surface functionalization by organic molecules, which for our purpose in this project are nontoxic and food-safe diethylenetriamine (DETA) and oligomeric polyethylenimine (PEI), thus DETA-CDots (manuscript in preparation) and PEI-CDots (more established CDots platform with the further progress in this project), respectively. However, CDots-like samples more popular in the literature are those obtained by thermal carbonization of organic precursors, with the potential of easier scale-up for our applications. We have further investigated the relationship between classical CDots and carbonization produced dot samples. On the latter our added innovation is to include specifically prepared CNPs in the precursor mixtures for carbonization, with excellent outcomes. We have made significant progress in our modified carbonization syntheses for dot samples in large quantities. We have incorporated the new findings and understandings from our investigations into our recent article (Small Structures 2025, 6, 2400455) to define more accurately the similarities and differences between classical CDots and carbonization produced dot samples. For the same purpose we used our specifically acquired experimental results to present a representative example in our recent paper published in the journal Micro (2025, 5, 14). 2. Evaluations and Comparisons of Dot Samples for Specific Project Needs. We have evaluated both classical CDots and carbonization produced dot samples (denoted as "nano-carbon/organic hybrids" or simply "hybrids"), especially their antibacterial functions comparatively. Our focus has been on DETA-CDots versus the carbonization produced hybrids denoted as DETA/CA and DETA/(CA+CNPs) (CA = citric acid), and similarly PEI-CDots versus PEI/CA and PEI/(CA+CNPs) hybrids. Our results from microscopy characterizations suggest composite-like structures in the hybrids, namely the sample structure of nano-carbons dispersed in crosslinked organic matters from the precursor mixtures, clearly different from the individually dispersed classical CDots. However, since the sample structural differences are at much shorter length scale than visible light wavelengths, optical spectroscopy can not differentiate the differences. Indeed, our results show that the absorption and fluorescence properties between the different dot samples are similar, including their similar fluorescence quantum yields, which according to our previously published studies (RSC Adv. 2017, 7, 30177) should be correlated with the antibacterial activities. We have evaluated the antibacterial functions of the different dot samples for various comparisons to identify those more suitable for our specific needs in antibacterial self-cleaning. Among the selected bacterial targets are Bacillus subtilis, Listeria monocytogenes, and Enterococcus faecalis. We have acquired a large amount of experimental data, some of which are still somewhat preliminary. Among the more conclusive results are those of the classical DETA-CDots against the drug-resistant L. monocytogenes, showing that the dot sample in 0.1 mg/mL solution coupled with one hour visible light exposure could kill up to 7 log of the Listeria cells. With the same experimental parameters and conditions DETA-CDots are also considerably effective in inactivating other bacterial cells including B. subtilis and the drug-resistant E. faecalis. Similarly, the classical PEI-CDots coupled with the visible light exposure are most effective in killing foodborne pathogenic L. monocytogenes cells, further confirming what we have previously reported on the potent antibacterial activities of PEI-CDots. In comparison we have found that in the solution phase the hybrid dot samples DETA/CA, DETA/(CA+CNPs), PEI/CA, and PEI/(CA+CNPs) produced via the thermal carbonization of the corresponding precursor mixtures are significantly less effective in the light-activated antibacterial activities. The findings should mostly be as expected for their being consistent with what we have previously found and reported (C 2022, 8, 54; DOI: 10.3390/c8040054) in terms of the mechanistic rationale that the composite-like hybrid dot samples in solutions are disadvantaged or limited by their slow diffusions in water to achieve the required close-neighbor interactions with the targeted bacterial cells within their photoexcited state lifetimes. However, for the purpose of antibacterial self-cleaning in this project, the application configurations should require no or very limited diffusions of the dot samples, which would suggest that the hybrid dot samples could still be a competitive alternative to classical CDots. 3. Design, Fabrication, and Antibacterial Evaluations of Polymer/Dot-Sample Composite Gel Formulations. Building upon the year-1 effort on the development of CDots-derived coating formulations with selected polymers as binders, we have further evaluated and validated the use of poly(vinyl alcohol) (PVA) as an excellent polymer binder or matrix for its benign and nontoxic nature, with the special property of being soluble in hot water but not so much in cold water, and its capacity to form both hydrogels and optically transparent films. We have designed and prepared formulations of PVA gels in which the selected dot samples are dispersed for the purpose of using these composite gel formulations as a facile and more controllable intermediate platform for antibacterial evaluations, toward the similar preparation and evaluations of the formulations in polymeric composite films and related configurations. Among the selected dot samples in the composite gel formulations are the comparable pairs of DETA-CDots versus DETA/CA and DETA/(CA+CNPs) and the pairs of PEI-CDots versus PEI/CA and PEI/(CA+CNPs). All these dot samples are readily miscible with PVA polymers in aqueous solutions. The resulting aqueous mixtures are then concentrated for the colored PVA composite blends/gels in which the dot samples are dispersed. For the antibacterial evaluations, clean microplate wells are coated with the composite gel formulations or with the corresponding blank PVA polymer gels as controls. The results obtained so far are generally very promising, even though some of the results may still be somewhat preliminary and require further confirmations. Specifically on the comparing pairs of DETA-CDots versus DETA/CA and DETA/(CA+CNPs) in the PVA composite gel formulations, they are all capable of inhibiting the growth of L. monocytogenes cells, an outcome clearly different from that of the pairs in the solution phase discussed in 2. above. The different outcomes are consistent with our rationale on the different diffusion characteristics of the different dot samples in solutions and also with our expectation that the carbonization produced dot samples in composite gels could be competitive in antibacterial activities. The comparing pairs of PEI-CDots versus PEI/CA and PEI/(CA+CNPs) in the PVA composite gels are all extremely effective in their antibacterial activities, killing both gram-positive L. monocytogenes and gram-negative S. typhimurium cells, thus much more potent than the DETA based/derived dot samples. The observed extreme effectiveness is obviously an extremely valuable outcome for our targeted self-cleaning applications. Next we will pursue more systematic antibacterial evaluations of these highly potent dot samples and their related sample configurations, including also investigations on the mechanistic origins and implications of the observed antibacterial properties, so as to establish the benchmark dot samples of benchmark performances for our project tasks in Aim 3.

Publications

• Type: Peer Reviewed Journal Articles Status: Published Year Published: 2025 Citation: Collins, J.; Yang, L.; Dong, X.; Sun, Y.-P. Antimicrobial Properties of Carbon Quantum Dots for Food Safety Applications. J. Nanoparticle Res., 2025, 27(2), article #35. (10.1007/s11051-025-06239-9)
• Type: Peer Reviewed Journal Articles Status: Published Year Published: 2025 Citation: Sun, Y.-P.; Yang, L. Carbon Dots from Preexisting Nanoparticles versus Carbonization  Similarity and Difference in Sample Structure-Morphology with Property and Mechanistic Consequences. Small Structures 2025, 6(3), 2400455. (10.1002/sstr.202400455)
• Type: Peer Reviewed Journal Articles Status: Published Year Published: 2025 Citation: Singh, B.; Adcock, A. F.; Dumra, S.; Collins, J.; Yang, L.; Bunker, C. E.; Qian, H.; Meziani, M. J.; Sun, Y.-P. Microwave-Assisted Carbonization Processing for Carbon Dots-Like Nanomaterials of Antimicrobial Properties. Micro 2025, 5, 14. (10.3390/micro5010014)
• Type: Peer Reviewed Journal Articles Status: Published Year Published: 2025 Citation: Sheriff, K.; Liang, W.; Yerra, S.; Sulejmanovic, D.; Nelson, C.; Lupini, A.; Hwu, S.-J.; Singh, B.; Sun, Y.-P. Carbon Dots from Surface-Capping/Passivation of Small Carbon Nanoparticles with Nanoscale Titanium Dioxide. Chem. Phys. Lett. 2025, 864, 141913. (10.1016/j.cplett.2025.141913)

Progress 08/15/23 to 08/14/24

Outputs
Target Audience:Scientists, professionals, and technical personals in the nanotech for food preservation and safety research community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?We have made major effort on providing hands-on laboratory research trainings for students and all team members in a highly interdisciplinary and cross-institutional project environment. The students at graduate and undergraduate levels have benefited significantly from their participation in the project. Especially encouraging has been that one participating graduate student graduated with PhD and took a R&D job in relevant industry and several participating undergraduate students went on to pursue their graduate education, which set up excellent examples for other participating students to look forward to similar opportunities in relevant technical fields. How have the results been disseminated to communities of interest?The research activities in this project have become a critical part of our overall effort in the development of carbon dots and their derived nanotechnology. We have been actively disseminating the results from our effort. The peer-reviewed journal publications acknowledging USDA support via this project are as follows. Liang, W.; Cao, L.; Scorzari, A.; McGrath, H.; Bunker, C. E.; Ren, X.; Wang, P.; Yang, L.; Sun, Y.-P. "Photoexcited State Properties of N-Ethylcarbazole-Functionalized Carbon Dots in Solution and in PVK Polymer Matrix" Chem. Phys. Lett. 2023, 833, 140964. (10.1016/j.cplett.2023.140964) Dong, X.; Liu, Y.; Adcock, A. F.; Sheriff, K.; Liang, W.; Yang, L.; Sun, Y.-P. "Carbon-TiO2 Hybrid Quantum Dots for Photocatalytic Inactivation of Gram-Positive and Gram-Negative Bacteria" Int. J. Mol. Sci. 2024, 25, 2196. (10.3390/ijms25042196) Liang, W.; Sheriff, K.; Singh, B.; Qian, H.; Dumra, S.; Collins, J.; Yang, L.; Sun, Y.-P. "On Carbon "Replacing" the Core in Classical Core/ZnS Quantum Dots" General Chem. 2024, 10 (1-2), 240001. (10.21127/yaoyigc20240001) What do you plan to do during the next reporting period to accomplish the goals?We have had a successful start of our project, which is clearly on the right track, so we will continue and expand our progress in the performance of the project tasks. In the next reporting period, we will still emphasize more on the tasks in Aim 1 and some tasks in Aim 2, but also initiate explorations on other project tasks, including those in Aim 3. We will disseminate as much as possible the results from our project to the nanotech for food preservation and safety research community through peer-reviewed publications and conference presentations. Of equal importance is our continuing focus on providing our graduate and undergraduate students with excellent trainings in the related research areas.

Impacts
What was accomplished under these goals? In the year-1 effort, we have successfully put together collaborative project teams at both partner institutions, with the teams composed of graduate students, research scientist, and undergraduate researchers. Our focus has been on project tasks for mostly Aim 1 and also Aim 2. The significant progress made in the performance of the project tasks is highlighted as follows. 1. Significant Progress in Food-Safe Carbon Dots Platform. Carbon dots (CDots) are classically defined as small carbon nanoparticles (CNPs) with effective surface passivation, which has been accomplished more successfully via the CNPs' surface functionalization with organic molecules. For food related applications of CDots, the use of food-safe organic molecules for the functionalization is obviously required or preferred. We have previously found and demonstrated that CDots with the small diamine molecule 2,2'-(ethylenedioxy)bis(ethylamine) (EDA) for the surface functionalization, thus EDA-CDots, are effective in the inactivation of bacterial pathogens, including multidrug-resistant ones. Molecular structure- and property-wise similar to EDA is the amino molecule diethylenetriamine (DETA), whose food-safe nature is reflected by its being on the FDA list of acceptable food additives, thus selected in this project for the functionalization of CNPs for DETA-CDots. We have made significant progress in the synthetic effort on DETA-CDots for the aim to have their antibacterial performance catch that of the more established EDA-CDots. The available results from antibacterial evaluations of DETA-CDots are very promising. For example, one tested sample of DETA-CDots at the concentration of 100 microgram nano-carbon (in the core CNPs of the CDots) per mL coupled with 1 hour of visible light exposure could kill all (>7 logs) Listeria monocytogenes (10403s) cells. We are pursuing further improvement in the synthesis of DETA-CDots for their being established as a benchmark food-safe CDots platform to be used in Aim 2 and Aim 3 tasks. 2. Dot Samples Tailored to Specific Project Needs. On the other CDots platform selected for this project, PEI-CDots (PEI = polyethylenimine oligomers), which is more established due to our previously more extensive investigations, our effort has been on the more efficient preparation/production of dot samples for the anticipated needs in other project tasks. In terms of the objectives of this project, the primary configuration with the use of CDots is for the dots to be on surface, loosely defined in such a way that the dots should be sufficiently exposed for interactions with bacteria species in the fluid phase but there must be no leaching into the fluid at all. For such a dual requirement, acceptable or even advantageous dot samples are those in which classical CDots or CDots-equivalent/like entities are dispersed in and crosslinked with some organic species, namely some specific versions of the "nano-carbon/organic hybrids" as we have defined in our recent publications. Such dot samples can be prepared efficiently by the thermal carbonization of selected organic precursors, including mixtures of PEI with citric acid (PEI/CA) for dot samples that are comparable with PEI-CDots for the specific needs in this project. In our year-1 effort, we have extensively and systematically investigated the carbonization of PEI/CA in different compositions and with varying processing conditions. Our results show that the formed nano-carbons (CNPs-like entities in the dot samples) are sourced not only from the carbonization of CA (commonly considered as being more readily carbonized) but also from PEI significantly, among other interesting and unexpected findings. Antibacterial evaluations of the dot samples have also been performed, with the results suggesting effective inactivation of several bacterial species. A manuscript reporting the synthesis, characterization, properties (including antibacterial results) of the dot samples is in the advanced stage of preparation. More interesting and potentially very significant and valuable are the results from the same carbonization synthesis with added preexisting CNPs, namely PEI/(CA+CNPs) as precursors for thermal carbonization processing. The dot samples thus prepared could have much higher nano-carbon contents, valuable in terms of high efficiency in the sample preparation and also other desirable sample properties that are not available in the absence of the added CNPs and in the classically synthesized PEI-CDots. Comprehensive characterizations of the dot samples prepared with different PEI/(CA+CNPs) precursor mixtures under various processing conditions are being pursued, so are the antibacterial evaluations of the selected dot samples. 3. Valuable Exploration of Antibacterial Coating Formulations. On the development of CDots-derived coating formulations with selected polymers as binders, which is a major project task for Aim 2, we have used aqueous compatible polymers poly(propionylethylenimine) (PPEI) and poly(vinyl alcohol) (PVA) to disperse PEI-CDots. PVA is particularly valuable for its characteristic property of no solubility in ambient water but soluble in hot water (85 C, for example) yet without precipitation in the subsequent cooling back to ambient temperature, thus ideally suited for coatings/films that are prepared in aqueous media and still remain intact for uses in aqueous media. As an important initial step, we have prepared thick gel-like formulations of PEI-CDots dispersed in polymeric PPEI and PVA, denoted as PPEI/PEI-CDots and PVA/PEI-CDots gels, respectively. We have used the gels in microplate wells for antibacterial tests (against Listeria monocytogenes as target, for example), from which the results have shown highly effective killing of the bacteria cells under visible light exposure conditions comparable with those used in similar evaluations of PEI-CDots in solution. However, interestingly and rather puzzlingly has been the observation of the similar killing without the light exposure, suggesting other antibacterial mechanisms at work. We are expanding the antibacterial evaluations for improved understandings.

Publications

• Type: Journal Articles Status: Published Year Published: 2024 Citation: Dong, X.; Liu, Y.; Adcock, A. F.; Sheriff, K.; Liang, W.; Yang, L.; Sun, Y.-P. CarbonTiO2 Hybrid Quantum Dots for Photocatalytic Inactivation of Gram-Positive and Gram-Negative Bacteria Int. J. Mol. Sci. 2024, 25, 2196. (10.3390/ijms25042196)
• Type: Journal Articles Status: Published Year Published: 2024 Citation: Liang, W.; Sheriff, K.; Singh, B.; Qian, H.; Dumra, S.; Collins, J.; Yang, L.; Sun, Y.-P. On Carbon Replacing the Core in Classical Core/ZnS Quantum Dots General Chem. 2024, 10 (1-2), 240001. (10.21127/yaoyigc20240001)
• Type: Journal Articles Status: Published Year Published: 2023 Citation: Liang, W.; Cao, L.; Scorzari, A.; McGrath, H.; Bunker, C. E.; Ren, X.; Wang, P.; Yang, L.; Sun, Y.-P. Photoexcited State Properties of N-Ethylcarbazole-Functionalized Carbon Dots in Solution and in PVK Polymer Matrix Chem. Phys. Lett. 2023, 833, 140964. (10.1016/j.cplett.2023.140964)