Aqueous exfoliation and dispersion of monolayer and bilayer graphene from graphite using sulfated cellulose nanofibrils.

Amphiphilic sulfated cellulose nanofibrils were synthesized with yields in excess of 99% by sulfation of dissolving pulp cellulose using chlorosulfonic acid in anhydrous N,N-dimethyl formamide followed by high-speed blending. The sulfation level was stoichiometrically tunable to between 1.48 and 2.23 mmol g-1. The optimized SCNF demonstrated the ability to act as an effective dispersant for graphene produced via exfoliation in aqueous media, allowing for the production of aqueous stabilized graphene with 3.9 ± 0.3 wt% graphite to graphene conversion and suspended solids comprised of 19.5 ± 1.5 wt% graphene. Graphene exfoliated with SCNF was observed to consist exclusively of mono- and bilayers, with 42% of sheets being monolayer. Furthermore, it was demonstrated that SCNF defibrillation and graphene exfoliation could be combined into a single one-pot process.

Regioselectively Carboxylated Cellulose Nanofibril Models from Dissolving Pulp: C6 via TEMPO Oxidation and C2,C3 via Periodate-Chlorite Oxidation.

Regioselective C6 and C2,C3 carboxylated cellulose nanofibrils (CNFs) have been robustly generated from dissolving pulp, a readily available source of unmodified cellulose, via stoichiometrically optimized 2,2,6,6-tetramethylpyperidine-1-oxyl (TEMPO)-mediated and sequential sodium periodate-sodium chlorite (PC) oxidation coupled with high-speed blending. Both regioselectively optimized carboxylated CNF series possess the widest ranges of comparable charges (0.72-1.48 mmol/g for T-CNFs vs. 0.72-1.10 mmol/g for PC-CNFs), but similar ranges of thickness (1.3-2.4 nm for T-CNF, 1.8-2.7 nm PC-CNF), widths (4.6-6.6 nm T-CNF, 5.5-5.9 nm PC-CNF), and lengths (254-481 nm T-CNF, 247-442 nm PC-CNF). TEMPO-mediated oxidation is milder and one-pot, thus more time and process efficient, whereas the sequential periodate-chlorite oxidation produces C2,C3 dialdehyde intermediates that are amenable to further chemical functionalization or post-reactions. These two well-characterized regioselectively carboxylated CNF series represent coherent cellulose nanomaterial models from a single woody source and have served as references for their safety study toward the development of a safer-by-design substance evaluation tool.

Life-cycle risk assessment of graphene-enabled textiles in fire protection gear.

A nanomaterial life-cycle risk assessment (Nano LCRA) of a graphene-enabled textile used in the construction of heat and fire-resistant personal protective equipment (PPE) was conducted to develop, analyze, and prioritize potential occupational, health and environmental risks. The analysis identifies potential receptors and exposure pathways at each product life-cycle stage and makes a qualitative evaluation of the potential significance of each scenario. A literature review, quality evaluation, and database were developed as part of the LCRA to identify potential hazards associated with graphene-based materials (GBMs) throughout the product life-cycle. Generally, risks identified from graphene-enabled textiles were low. Of the developed exposure scenarios, occupational inhalation exposures during raw material and product manufacturing ranked highest. The analysis identifies the key potential human and environmental hazards and exposures of the products across the product life-cycle of graphene enabled textiles. Priority research gaps to reduce uncertainty include evaluating long-term, low dose graphene exposures typical of the workplace, as well as the potential release and hazard characterization of graphene-acrylic nanocomposites.

Cultured Meat Safety Research Priorities: Regulatory and Governmental Perspectives.

As with every new technology, safety demonstration is a critical component of bringing products to market and gaining public acceptance for cultured meat and seafood. This manuscript develops research priorities from the findings of a series of interviews and workshops with governmental scientists and regulators from food safety agencies in fifteen jurisdictions globally. The interviews and workshops aimed to identify the key safety questions and priority areas of research. Participants raised questions about which aspects of cultured meat and seafood production are novel, and the implications of the paucity of public information on the topic. Novel parameters and targets may require the development of new analytical methods or adaptation and validation of existing ones, including for a diversity of product types and processes. Participants emphasized that data sharing of these efforts would be valuable, similar to those already developed and used in the food and pharmaceutical fields. Contributions to such databases from the private and public sectors would speed general understanding as well as efforts to make evaluations more efficient. In turn, these resources, combined with transparent risk assessment, will be critical elements of building consumer trust in cultured meat and seafood products.

Review on Hybrid Reinforced Polymer Matrix Composites with Nanocellulose, Nanomaterials, and Other Fibers.

The use of composite materials has seen many new innovations for a large variety of applications. The area of reinforcement in composites is also rapidly evolving with many new discoveries, including the use of hybrid fibers, sustainable materials, and nanocellulose. In this review, studies on hybrid fiber reinforcement, the use of nanocellulose, the use of nanocellulose in hybrid forms, the use of nanocellulose with other nanomaterials, the applications of these materials, and finally, the challenges and opportunities (including safety issues) of their use are thoroughly discussed. This review will point out new prospects for the composite materials world, enabling the use of nano- and micron-sized materials together and creating value-added products at the industrial scale. Furthermore, the use of hybrid structures consisting of two different nano-materials creates many novel solutions for applications in electronics and sensors.

Food safety considerations and research priorities for the cultured meat and seafood industry.

Cell-cultured meat and seafood offer a sustainable opportunity to meet the world's increasing demand for protein in a climate-changed world. A responsible, data-driven approach to assess and demonstrate safety of cell-cultured meat and seafood can support consumer acceptance and help fully realize the potential of these products. As an initial step toward a thorough demonstration of safety, this review identifies hazards that could be introduced during manufacturing, evaluates applicability of existing safety assessment approaches, and highlights research priorities that could support safe commercialization. Input was gathered from members of the cultured meat and seafood industry, researchers, regulators, and food safety experts. A series of workshops were held with 87 industry representatives and researchers to create a modular manufacturing process diagram, which served as a framework to identify potential chemical and biological hazards along the steps of the manufacturing process that could affect the safety of a final food product. Interviews and feedback on draft documents validated the process diagram and supported hazard identification and evaluation of applicable safety methods. Most hazards are not expected to be novel; therefore, safety assessment methods from a range of fields, such as conventional and novel foods, foods produced from biotechnology, pharmaceuticals, and so forth, are likely to be applicable. However, additional assessment of novel inputs or products with significant differences from existing foods may be necessary. Further research on the safety of the inputs and associated residues, potential for contamination, and development of standardized safety assessment approaches (particularly animal-free methods) is recommended.

Fluorescently Labeled Cellulose Nanofibers for Environmental Health and Safety Studies.

An optimal methodology for locating and tracking cellulose nanofibers (CNFs) in vitro and in vivo is crucial to evaluate the environmental health and safety properties of these nanomaterials. Here, we report the use of a new boron-dipyrromethene (BODIPY) reactive fluorescent probe, meso-DichlorotriazineEthyl BODIPY (mDTEB), tailor-made for labeling CNFs used in simulated or in vivo ingestion exposure studies. Time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) was used to confirm covalent attachment and purity of mDTEB-labeled CNFs. The photoluminescence properties of mDTEB-labeled CNFs, characterized using fluorescence spectroscopy, include excellent stability over a wide pH range (pH2 to pH10) and high quantum yield, which provides detection at low (µM) concentrations. FLIM analysis also showed that lignin-like impurities present on the CNF reduce the fluorescence of the mDTEB-labeled CNF, via quenching. Therefore, the chemical composition and the methods of CNF production affect subsequent studies. An in vitro triculture, small intestinal, epithelial model was used to assess the toxicity of ingested mDTEB-labeled CNFs. Zebrafish (Danio rerio) were used to assess in vivo environmental toxicity studies. No cytotoxicity was observed for CNFs, or mDTEB-labeled CNFs, either in the triculture cells or in the zebrafish embryos.

A methodology for developing key events to advance nanomaterial-relevant adverse outcome pathways to inform risk assessment.

Significant advances have been made in the development of Adverse Outcome Pathways (AOPs) over the last decade, mainly focused on the toxicity mechanisms of chemicals. These AOPs, although relevant to manufactured nanomaterials (MNs), do not currently capture the reported roles of size-associated properties of MNs on toxicity. Moreover, some AOs of relevance to airborne exposures to MNs such as lung inflammation and fibrosis shown in animal studies may not be targeted in routine regulatory decision making. The primary objective of the present study was to establish an approach to advance the development of AOPs of relevance to MNs using existing, publicly available, nanotoxicology literature. A systematic methodology was created for curating, organizing and applying the available literature for identifying key events (KEs). Using a case study approach, the study applied the available literature to build the biological plausibility for 'tissue injury', a KE of regulatory relevance to MNs. The results of the analysis reveal the various endpoints, assays and specific biological markers used for assessing and reporting tissue injury. The study elaborates on the limitations and opportunities of the current nanotoxicology literature and provides recommendations for the future reporting of nanotoxicology results that will expedite not only the development of AOPs for MNs but also aid in application of existing data for decision making.

Translating Scientific Advances in the AOP Framework to Decision Making for Nanomaterials.

Much of the current innovation in advanced materials is occurring at the nanoscale, specifically in manufactured nanomaterials (MNs). MNs display unique attributes and behaviors, and may be biologically and physically unique, making them valuable across a wide range of applications. However, as the number, diversity and complexity of MNs coming to market continue to grow, assessing their health and environmental risks with traditional animal testing approaches is too time- and cost-intensive to be practical, and is undesirable for ethical reasons. New approaches are needed that meet current requirements for regulatory risk assessment while reducing reliance on animal testing and enabling safer-by-design product development strategies to be implemented. The adverse outcome pathway (AOP) framework presents a sound model for the advancement of MN decision making. Yet, there are currently gaps in technical and policy aspects of AOPs that hinder the adoption and use for MN risk assessment and regulatory decision making. This review outlines the current status and next steps for the development and use of the AOP framework in decision making regarding the safety of MNs. Opportunities and challenges are identified concerning the advancement and adoption of AOPs as part of an integrated approach to testing and assessing (IATA) MNs, as are specific actions proposed to advance the development, use and acceptance of the AOP framework and associated testing strategies for MN risk assessment and decision making. The intention of this review is to reflect the views of a diversity of stakeholders including experts, researchers, policymakers, regulators, risk assessors and industry representatives on the current status, needs and requirements to facilitate the future use of AOPs in MN risk assessment. It incorporates the views and feedback of experts that participated in two workshops hosted as part of an Organization for Economic Cooperation and Development (OECD) Working Party on Manufactured Nanomaterials (WPMN) project titled, "Advancing AOP Development for Nanomaterial Risk Assessment and Categorization", as well as input from several EU-funded nanosafety research consortia.

Physical, chemical, and toxicological characterization of fibrillated forms of cellulose using an in vitro gastrointestinal digestion and co-culture model.

Fibrillated cellulose is a next-generation material in development for a variety of applications, including use in food and food-contact materials. An alternative testing strategy including simulated digestion was developed to compare the physical, chemical, and biological characteristics of seven different types of fibrillated cellulose, following European Food Safety Authority guidance. Fibrillated forms were compared to a conventional form of cellulose which has been used in food for over 85 years and has Generally Recognized as safe regulatory status in the USA. The physical and chemical characterization of fibrillated celluloses demonstrate that these materials are similar physically and chemically, which composed of the same fundamental molecular structure and exhibit similar morphology, size, size distribution, surface charge, and low levels of impurities. Simulated gastrointestinal and lysosomal digestions demonstrate that these physical and chemical similarities remain following exposure to conditions that mimic the gastrointestinal tract or intracellular lysosomes. A toxicological investigation with an advanced intestinal co-culture model found that exposure to each of the fibrillated and conventional forms of cellulose, in either the pristine or digested form at 0.4% by weight, showed no adverse toxicological effects including cytotoxicity, barrier integrity, oxidative stress, or inflammation. The results demonstrate the physical, chemical, and biological similarities of these materials and provide substantive evidence to support their grouping and ability to read-across data as part of a food safety demonstration.

A 90-day dietary study with fibrillated cellulose in Sprague-Dawley rats.

Novel forms of fibrillated cellulose offer improved attributes for use in foods. Conventional cellulose and many of its derivatives are already widely used as food additives and are authorized as safe for use in foods in many countries. However, novel forms have not yet been thoroughly investigated using standardized testing methods. This study assesses the 90-day dietary toxicity of fibrillated cellulose, as compared to a conventional cellulose, Solka Floc. Sprague Dawley rats were fed 2 %, 3 %, or 4 % fibrillated cellulose for 90 consecutive days, and parallel Solka Floc groups were used as controls. Survival, clinical observations, body weight, food consumption, ophthalmologic evaluations, hematology, serum chemistry, urinalysis, post-mortem anatomic pathology, and histopathology were monitored and performed. No adverse observations were noted in relation to the administration of fibrillated cellulose. Under the conditions of this study and based on the toxicological endpoints evaluated, the no-observed-adverse-effect level (NOAEL) for fibrillated cellulose was 2194.2 mg/kg/day (males) and 2666.6 mg/kg/day (females), corresponding to the highest dose tested (4 %) for male and female Sprague Dawley rats. These results demonstrate that fibrillated cellulose behaves similarly to conventional cellulose and raises no safety concerns when used as a food ingredient at these concentrations.

Physical, chemical, and toxicological characterization of sulfated cellulose nanocrystals for food-related applications using in vivo and in vitro strategies.

Cellulose nanocrystals (CNCs) are a next-generation cellulose product with many unique properties including applications in the food industry as a food additive, food coating, and in food-contact packaging material. While CNC is anticipated to be safe due to its similarity to the many forms of cellulose currently used as food additives, special consideration is given to it as it is the first manufactured form of cellulose that is nanoscale in both length and width. A proactive approach to safety has been adopted by manufacturers to demonstrate CNC safety toward responsible commercialization. As part of the safety demonstration, in vivo and in vitro testing strategies were commissioned side-by-side with conventional cellulose, which has been safely used in food for decades. Testing included a 90-day rodent feeding study as well as additional physical, chemical, and biological studies in vitro that follow European Food Safety Authority (EFSA) guidance to demonstrate the safe use of novel food ingredients. The strategy includes assessment of neat materials side-by-side with simulated digestion, mimicking conditions that occur along the gastrointestinal tract as well as intracellularly. An intestinal co-culture model examined any potential toxicological effects from exposure to either pristine or digested forms of CNC including cytotoxicity, metabolic activity, membrane permeability, oxidative stress, and proinflammatory responses. None of the studies demonstrated any toxicity via oral or simulated oral exposure. These studies demonstrate that CNC produced by InnoTech Alberta is similarly safe by ingestion as conventional cellulose with a no-observed-adverse-effect level of 2085.3 (males) and 2682.8 (females) mg/kg/day.

Risk Analysis of Cellulose Nanomaterials by Inhalation: Current State of Science.

Cellulose nanomaterials (CNs) are emerging advanced materials with many unique properties and growing commercial significance. A life-cycle risk assessment and environmental health and safety roadmap identified potential risks from inhalation of powdered CNs in the workplace as a key gap in our understanding of safety and recommended addressing this data gap to advance the safe and successful commercialization of these materials. Here, we (i) summarize the currently available published literature for its contribution to our current understanding of CN inhalation hazard and (ii) evaluate the quality of the studies for risk assessment purposes using published study evaluation tools for nanomaterials to assess the weight of evidence provided. Our analysis found that the quality of the available studies is generally inadequate for risk assessment purposes but is improving over time. There have been some advances in knowledge about the effects of short-term inhalation exposures of CN. The most recent in vivo studies suggest that short-term exposure to CNs results in transient inflammation, similarly to other poorly soluble, low toxicity dusts such as conventional cellulose, but is markedly different from fibers with known toxicity such as certain types of multiwalled carbon nanotubes or asbestos. However, several data gaps remain, and there is still a lack of understanding of the effects from long-term, low-dose exposures that represent realistic workplace conditions, essential for a quantitative assessment of potential health risk. Therefore, taking precautions when handling dry forms of CNs to avoid dust inhalation exposure is warranted.

Evaluating mucoadhesion properties of three types of nanocellulose in the gastrointestinal tract in vitro and ex vivo.

The mucoadhesive properties of three types of nanocellulose (CNF, CNC, and Tempo-CNF) was investigated in the digestive condition with ex vivo and in vitro assays. In the ex vivo flow-through method, three nanocellulose materials showed different levels of retention on porcine gastric and intestinal mucosal surfaces. Fluorescence microscopy confirmed that retention of CNF could be due to entanglement with the mucosal layer, while retention of Tempo-CNF could be due to instantaneous gelling on the mucosal surface. In an in vitro viscometric method, 2% CNC showed the highest viscosity synergism (relative enhancement=11.80 ± 1.14) in the gastric condition, while Tempo-CNF only displayed synergism under gelling concentrations (0.1%). Evaluation of zeta potential revealed that 0.025-0.1% CNC interacted with mucin particles by changing the surface charge of the mucin-nanocellulose system. These results indicate that nanocellulose shows mucoadhesive properties in digestive tract, where the level of adhesion depends on type of nanocellulose, its concentration and the gastrointestinal section.

Current characterization methods for cellulose nanomaterials.

A new family of materials comprised of cellulose, cellulose nanomaterials (CNMs), having properties and functionalities distinct from molecular cellulose and wood pulp, is being developed for applications that were once thought impossible for cellulosic materials. Commercialization, paralleled by research in this field, is fueled by the unique combination of characteristics, such as high on-axis stiffness, sustainability, scalability, and mechanical reinforcement of a wide variety of materials, leading to their utility across a broad spectrum of high-performance material applications. However, with this exponential growth in interest/activity, the development of measurement protocols necessary for consistent, reliable and accurate materials characterization has been outpaced. These protocols, developed in the broader research community, are critical for the advancement in understanding, process optimization, and utilization of CNMs in materials development. This review establishes detailed best practices, methods and techniques for characterizing CNM particle morphology, surface chemistry, surface charge, purity, crystallinity, rheological properties, mechanical properties, and toxicity for two distinct forms of CNMs: cellulose nanocrystals and cellulose nanofibrils.

Comment on Shvedova et al. (2016), "gender differences in murine pulmonary responses elicited by cellulose nanocrystals".

A recent publication in "Particle and Fibre Toxicology" reported on the gender differences in pulmonary toxicity from oro-pharyngeal aspiration of a high dose of cellulose nanocrystals. The study is timely given the growing interest in diverse commercial applications of cellulose nanomaterials, and the need for studies addressing pulmonary toxicity. The results from this study are interesting and can be strengthened with a discussion of how differences in the weights of female and male C57BL/6 mice was accounted for. Without such a discussion, the observed differences could be partially explained by the lower body weights of females, resulting in higher doses than males when standardized to body weight or lung volume. Further, few conclusions can be drawn about the pulmonary toxicity of cellulose nanocrystals given the study design: examination of a single high dose of cellulose nanocrystals, administered as a bolus, without positive or negative controls or low dose comparisons, and at an unphysiological and high dose rate. Simulating the bolus type delivery by inhalation would require a highly unrealistic exposure concentration in the g/m3 range of extremely short duration. A discussion of these limitations is missing in the paper; further speculative comparisons of cellulose nanocrystals toxicity to asbestos and carbon nanotubes in the abstract are both unwarranted and can be misleading, these materials were neither mentioned in the manuscript, nor evaluated in the study.

Framework to Evaluate Exposure Relevance and Data Needs for Risk Assessment of Nanomaterials using in Vitro Testing Strategies.

This article presents a multistage framework for evaluating the strength of evidence of nanomaterial (NM) exposure characterization data to optimize the utility of in vitro testing strategies for human health risk assessment. This framework is intended to aid risk assessors in evaluating the relevance of data from in vitro tests and to optimize the development of new in vitro testing strategies. The initial stage frames the exposure scenarios of interest in advance of testing to incorporate aspects such as release points, route of exposure, biological and environmental transformations, dose metrics, and biological targets in subsequent stages. The second stage considers characterization in the context of a realistic exposure and the third stage involves designing a testing strategy based on expected exposure conditions. For the fourth and final stage, we propose a matrix approach to evaluate the strength of evidence obtained in the first three stages as a basis for determining the best combination of test conditions and analytical methods available to characterize and measure exposure based on the NM type. This approach can also be used to evaluate existing data for their relevance to the expected exposure scenario and to further develop and optimize in vitro testing strategies. Implementation of the proposed strategy will generate meaningful information on NM properties and their interaction with biological systems, based on realistic exposure scenarios, which will be cost effective and can be applied for assessing risk and making intelligent regulatory decisions regarding the use and disposal of NMs.

The Role of the Food Matrix and Gastrointestinal Tract in the assessment of biological properties of ingested engineered nanomaterials (iENMs): State of the science and knowledge gaps.

Many foods contain appreciable levels of engineered nanomaterials (ENMs) (diameter < 100 nm) that may be either intentionally or unintentionally added. These ENMs vary considerably in their compositions, dimensions, morphologies, physicochemical properties, and biological responses. From a toxicological point of view, it is often convenient to classify ingested ENMs (iENMs) as being either inorganic (such as TiO2, SiO2, Fe2O3, or Ag) or organic (such as lipid, protein, or carbohydrate), since the former tend to be indigestible and the latter are generally digestible. At present there is a relatively poor understanding of how different types of iENMs behave within the human gastrointestinal tract (GIT), and how the food matrix and biopolymers transform their physico-chemical properties and influence their gastrointestinal fate. This lack of knowledge confounds an understanding of their potential harmful effects on human health. The purpose of this article is to review our current understanding of the GIT fate of iENMs, and to highlight gaps where further research is urgently needed in assessing potential risks and toxicological implications of iENMs. In particular, a strong emphasis is given to the development of standardized screening methods that can be used to rapidly and accurately assess the toxicological properties of iENMs.