Occurrence, bioaccumulation, fate, and risk assessment of emerging pollutants in aquatic environments: A review.

Significant concerns on a global scale have been raised in response to the potential adverse impacts of emerging pollutants (EPs) on aquatic creatures. We have carefully reviewed relevant research over the past 10 years. The study focuses on five typical EPs: pharmaceuticals and personal care products (PPCPs), per- and polyfluoroalkyl substances (PFASs), drinking water disinfection byproducts (DBPs), brominated flame retardants (BFRs), and microplastics (MPs). The presence of EPs in the global aquatic environment is source-dependent, with wastewater treatment plants being the main source of EPs. Multiple studies have consistently shown that the final destination of most EPs in the water environment is sludge and sediment. Simultaneously, a number of EPs, such as PFASs, MPs, and BFRs, have long-term environmental transport potential. Some EPs exhibit notable tendencies towards bioaccumulation and biomagnification, while others pose challenges in terms of their degradation within both biological and abiotic treatment processes. The results showed that, in most cases, the ecological risk of EPs in aquatic environments was low, possibly due to potential dilution and degradation. Future research topics should include adding EPs detection items for the aquatic environment, combining pollution, and updating prediction models.

A Systematic Review on Microplastic Contamination in Fishes of Asia: Polymeric Risk Assessment and Future Prospectives.

Microplastics (MPs) have attracted global concern because of their harmful effects on marine biota; their toxic properties can negatively impact aquatic ecosystems. Fish is an essential source of protein for humans, playing a crucial role in daily food intake. Until recently, MPs were addressed primarily as environmental pollutants, but they are now increasingly recognized as contaminants in the food supply. The present review has comprehended the current knowledge of MP contamination in freshwater and marine fishes of Asia, including 112 peer-reviewed sources from 2016 to 2023. The review recorded 422 Asian fishes (345 marine and 77 freshwater) to be contaminated with MPs. Clarias gariepinus and Selaroides leptolepi have shown maximum MP contamination in the freshwater and marine environments of Asia, respectively. Omnivorous and carnivorous fishes exhibited higher susceptibility to ingesting MPs. Benthopelagic, demersal, and reef-associated habitats were identified as more prone to MP accumulation. In both freshwater and marine environments, China has the highest number of contaminated species among all the countries. Pollution indices indicated high MP contamination in both freshwater and marine environments. A prevalence of fibers was recorded in all fishes. Black- and blue-colored MPs of <500 µm-1 mm size were found dominantly. Polyethylene terephthalate and polyethylene were recorded as the prevalent plastic polymers in freshwater and marine fish, respectively. Overall, the review served as a comprehensive understanding of MP concentrations and variations between species, between feeding habits, and between geographic locations, which can be pivotal for addressing pressing environmental challenges, protecting human health, and fostering global sustainability efforts in the face of escalating plastic pollution. Environ Toxicol Chem 2024;43:671-685. © 2024 SETAC.

Designing a circular carbon and plastics economy for a sustainable future.

The linear production and consumption of plastics today is unsustainable. It creates large amounts of unnecessary and mismanaged waste, pollution and carbon dioxide emissions, undermining global climate targets and the Sustainable Development Goals. This Perspective provides an integrated technological, economic and legal view on how to deliver a circular carbon and plastics economy that minimizes carbon dioxide emissions. Different pathways that maximize recirculation of carbon (dioxide) between plastics waste and feedstocks are outlined, including mechanical, chemical and biological recycling, and those involving the use of biomass and carbon dioxide. Four future scenarios are described, only one of which achieves sufficient greenhouse gas savings in line with global climate targets. Such a bold system change requires 50% reduction in future plastic demand, complete phase-out of fossil-derived plastics, 95% recycling rates of retrievable plastics and use of renewable energy. It is hard to overstate the challenge of achieving this goal. We therefore present a roadmap outlining the scale and timing of the economic and legal interventions that could possibly support this. Assessing the service lifespan and recoverability of plastic products, along with considerations of sufficiency and smart design, can moreover provide design principles to guide future manufacturing, use and disposal of plastics.

Assessment of Nonalcoholic Fatty Liver Disease Symptoms and Gut-Liver Axis Status in Zebrafish after Exposure to Polystyrene Microplastics and Oxytetracycline, Alone and in Combination.

BACKGROUND: Environmental pollution may give rise to the incidence and progression of nonalcoholic fatty liver disease (NAFLD), the most common cause for chronic severe liver lesions. Although knowledge of NAFLD pathogenesis is particularly important for the development of effective prevention, the relationship between NAFLD occurrence and exposure to emerging pollutants, such as microplastics (MPs) and antibiotic residues, awaits assessment. OBJECTIVES: This study aimed to evaluate the toxicity of MPs and antibiotic residues related to NAFLD occurrence using the zebrafish model species. METHODS: Taking common polystyrene MPs and oxytetracycline (OTC) as representatives, typical NAFLD symptoms, including lipid accumulation, liver inflammation, and hepatic oxidative stress, were screened after 28-d exposure to environmentally realistic concentrations of MPs (0.69mg/L) and antibiotic residue (3.00µg/L). The impacts of MPs and OTC on gut health, the gut-liver axis, and hepatic lipid metabolism were also investigated to reveal potential affecting mechanisms underpinning the NAFLD symptoms observed. RESULTS: Compared with the control fish, zebrafish exposed to MPs and OTC exhibited significantly higher levels of lipid accumulation, triglycerides, and cholesterol contents, as well as inflammation, in conjunction with oxidative stress in their livers. In addition, a markedly smaller proportion of Proteobacteria and higher ratios of Firmicutes/Bacteroidetes were detected by microbiome analysis of gut contents in treated samples. After the exposures, the zebrafish also experienced intestinal oxidative injury and yielded significantly fewer numbers of goblet cells. Markedly higher levels of the intestinal bacteria-sourced endotoxin lipopolysaccharide (LPS) were also detected in serum. Animals treated with MPs and OTC exhibited higher expression levels of LPS binding receptor (LBP) and downstream inflammation-related genes while also exhibiting lower activity and gene expression of lipase. Furthermore, MP-OTC coexposure generally exerted more severe effects compared with single MP or OTC exposure. DISCUSSION: Our results suggested that exposure to MPs and OTC may disrupt the gut-liver axis and be associated with NAFLD occurrence. https://doi.org/10.1289/EHP11600.

Cost effective media optimization for PHB production by Bacillus badius MTCC 13004 using the statistical approach.

Polyhydroxybutyrate (PHB) has significant potential for replacing non-biodegradable traditional plastic, which is responsible for several global environmental issues. The main problem with switching to bio-based alternatives for petrochemical plastics is the large price gap on the market. To overcome this problem, the present research was focused on the utilization of inexpensive substrates i.e. agricultural residues for cost-effective PHB production by endospore-forming bacteria Bacillus badius MTCC 13004. For efficient PHB production, Box-Behnken Design (BBD) was selected for media optimization and to observe the interactive effects of four variables i.e. pH, Na acetate, Banana peel, and mustard cake. PHB yield of 2.11 g/L was attained under optimized conditions compared to non-optimized conditions (0.72 g/L). FTIR spectra analysis of PHB extracted from Bacillus badius was found to be similar to commercial PHB. NMR data was also matched with the chemical shift signals CH, CH2, and CH3 of PHB. The melting temperature (Tm) and glass transition temperature (Tg) of PHB from Bacillus badius was found to be 165.14 and 2.68 °C, respectively. Further, PCR protocol was also designed to amplify key enzymes of the PHB synthesis pathway i.e. PHB synthase (phb C gene).

In vitro toxicity assessment of polyethylene terephthalate and polyvinyl chloride microplastics using three cell lines from rainbow trout (Oncorhynchus mykiss).

The RTgill-W1 (gill), RTG-2 (gonad), and RTL-W1 (liver) cell lines derived from a freshwater fish rainbow trout (Oncorhynchus mykiss), were used to assess the toxicity of polyethylene terephthalate (PET) and two forms of polyvinyl chloride (PVC). Two size fractions (25-µm and 90-µm particles) were tested for all materials. The highest tested concentration was 1 mg/ml, corresponding to from 70 000 ± 9000 to 620 000 ± 57 000 particles/ml for 25-µm particles and from 2300 ± 100 to 11 000 ± 1000 particles/ml for 90-µm particles (depending on the material). Toxicity differences between commercial PVC dry blend powder and secondary microplastics created from a processed PVC were newly described. After a 24-h exposure, the cells were analyzed for changes in viability, 7-ethoxyresorufin-O-deethylase (EROD) activity, and reactive oxygen species (ROS) generation. In addition to the microplastic suspensions, leachates and particles remaining after leaching resuspended in fresh exposure medium were tested. The particles were subjected to leaching for 1, 8, and 15 days. The PVC dry blend (25 µm and 90 µm) and processed PVC (25 µm) increased ROS generation, to which leached chemicals appeared to be the major contributor. PVC dry blend caused substantially higher ROS induction than processed PVC, showing that the former is not suitable for toxicity testing, as it can produce different results from those of secondary PVC. The 90-µm PVC dry blend increased ROS generation only after prolonged leaching. PET did not induce any changes in ROS generation, and none of the tested polymers had any effect on viability or EROD activity. The importance of choosing realistic extraction procedures for microplastic toxicity experiments was emphasized. Conducting long-term experiments is crucial to detect possible environmentally relevant effects. In conclusion, the tested materials showed no acute toxicity to the cell lines.

Assessment of microplastic bioconcentration, bioaccumulation and biomagnification in a simple coral reef food web.

Plastics, and more specifically, microplastics (MPs, <5 mm) are considered a marine contaminant of emerging concern. To accurately assess the ecological risk of MPs, it is critical to first understand the relationship between MP contamination in organisms with that in their surrounding environment. The goal of this study was to examine the ecological risk of MPs in coral reef ecosystems by assessing the MP contamination found within a simple food web against contamination in the surrounding environment. Taxa representing three trophic levels (zooplankton, benthic crustaceans, and reef fish), as well as the distinct environmental matrices which they inhabit (i.e., mid-column water and sediment) were collected from two mid-shelf reefs in the central Great Barrier Reef, Australia. Microplastics were isolated using validated clarification techniques, visually characterised (i.e., shape, colour, size) by microscopy, chemically confirmed by Fourier transform infrared spectroscopy and recorded in all three trophic levels and all abiotic samples. MPs were found to bioconcentrate, with similar concentrations, polymer types, sizes, shapes, and colours at each trophic level compared to their surrounding environment. However, MP contamination varied across the three trophic levels, with no evidence of bioaccumulation. Further, MP concentrations did not increase up the food web, discounting MP biomagnification. Regardless, given the heterogeneity of MPs found in the marine environment, and the complexity of marine food webs, trophic transfer represents a prominent pathway of exposure from lower to higher trophic levels.

Advanced oxidation processes for the elimination of microplastics from aqueous systems: Assessment of efficiency, perspectives and limitations.

Microplastics act as a vector of heavy metals, organic pollutants, pathogens and resistance genes in the environment further aggravate the pollution of plastics. The conventional wastewater/water treatment processes can physically capture and remove most of microplastics, but the success rates varies. How to quickly remove a large amount of microplastics from aqueous system is a key research topic at present. Recently, advanced oxidation processes (AOPs) as a green elimination strategy has attracted attention because of its effective elimination, strong destruction and safety. The molecular chain of microplastics can be gradually degraded into small molecular organics until H2O and CO2 by strong oxidizing free radical produced by AOPs. Unfortunately, problematically, the elimination of microplastics in aqueous system by AOPs is recently carried out on a laboratory scale. The application and implementation of this strategy are restricted by long reaction time, low liquid phase degradation efficiency and the formation of nanoplastics. Generally, the technology is still in its infancy, and most studies are carried out under laboratory conditions. The degradation of microplastics in aqueous system also needs appropriate conditions, but it is not always feasible under field conditions in AOPs. Although AOPs can be used as a green degradation technology to eliminate microplastics in aqueous systems in theory, it still needs to be furtherly explored in practical application. Consequently, before AOPs as a green elimination strategy is successfully applied to the effective remove microplastics, more in-depth research is still required, such as the setting from single condition to complex environment, the transfer from laboratory scale to field scale, and systematic toxicity evaluation of corresponding products.

Different patterns of hypoxia aggravate the toxicity of polystyrene nanoplastics in the mussels Mytilus galloprovincialis: Environmental risk assessment of plastics under global climate change.

Hypoxia, largely triggered by anthropogenic activities and global climate change, exerts widespread and expanding stress on marine ecosystems. As an emerging contaminant, the influence of nanoplastics on marine organisms has also attracted attention in recent years. However, the impact of hypoxia on the risk assessments of nanoplastics is rarely considered. This study investigated the toxicity of PS-NPs (0, 0.5, and 5 mg/L) to the coastal mussels Mytilus galloprovincialis under different patterns of hypoxia (normoxia, constant hypoxia, and fluctuating hypoxia). The results showed that constant hypoxia might reduce the accumulation of PS-NPs in mussels by decreasing the standard metabolic rate. The impairment of PS-NPs on mussel immunity was also exacerbated by constant hypoxia. Fluctuating hypoxia did not affect the accumulation of PS-NPs, but aggravated the oxidative damage caused by PS-NPs. These findings emphasize the importance of environmental factors and their temporal variability in plastic risk assessment.

Cyanobacterial Polyhydroxyalkanoates: A Sustainable Alternative in Circular Economy.

Conventional petrochemical plastics have become a serious environmental problem. Its unbridled use, especially in non-durable goods, has generated an accumulation of waste that is difficult to measure, threatening aquatic and terrestrial ecosystems. The replacement of these plastics with cleaner alternatives, such as polyhydroxyalkanoates (PHA), can only be achieved by cost reductions in the production of microbial bioplastics, in order to compete with the very low costs of fossil fuel plastics. The biggest costs are carbon sources and nutrients, which can be appeased with the use of photosynthetic organisms, such as cyanobacteria, that have a minimum requirement for nutrients, and also using agro-industrial waste, such as the livestock industry, which in turn benefits from the by-products of PHA biotechnological production, for example pigments and nutrients. Circular economy can help solve the current problems in the search for a sustainable production of bioplastic: reducing production costs, reusing waste, mitigating CO2, promoting bioremediation and making better use of cyanobacteria metabolites in different industries.

First occurrence and composition assessment of microplastics in native mussels collected from coastal and offshore areas of the northern and central Adriatic Sea.

In recent years, the occurrence of microplastics in the aquatic environment has gathered increasing scientific interest. Several studies have shown that the ingestion of microplastics may negatively influence the physiology of marine organisms having different feeding strategies, particularly in those species which cannot discriminate between food sources. Recent studies highlighted the potential for such particles to accumulate in the food web, posing risks to human health via the consumption of seafood. Furthermore, early findings also indicated the role of microplastics as vectors of chemical pollutants either used as additives during synthesis of the plastics or adsorbed directly from seawater, i.e., PAHs, PCB, and surfactants. Despite the importance of microplastics in adsorption and transport of hydrophobic pollutants, little is known about their distribution and accumulation in marine food webs, or their direct and indirect harmful effects. The Adriatic Sea represents a semi-enclosed basin with a low water recirculation rate and high anthropogenic pressures associated with unsustainable fishing and inputs of contaminants. The body burden, accumulation rates, polymer composition, and recurring morphotypes of microplastics in native blue mussels (M. galloprovincialis) were examined. Organisms collected offshore were compared to those collected in coastal areas. Microplastics were recovered from the soft tissues of all analyzed mussels. Coastal organisms showed a load of 1.06-1.33 fragments g-1 (wet weight) and 0.62-0.63 fibers g-1 (wet weight) while offshore organisms showed an accumulation of 0.65-0.66 fragments g-1 (wet weight) and 0.24-0.35 fibers g-1 (wet weight). The size class distribution revealed a marked prevalence of smaller particles (20 µm to 40 µm range) and the most recurring polymer type in analyzed organisms was PE followed by PP, PET, and equal amounts of PS, PLY, and PVC. A significant site-, time-, and oceanographic-related distribution trend was observed. Based on the findings presented here, there is a clear need to implement a seafood safety monitoring program to better understand actual human health-related risks.

Application of catastrophe theory in comprehensive ecological security assessment of plastic greenhouse soil contaminated by phthalate esters.

Large amount of phthalate esters (PAEs) used as plasticizers in polyvinyl chloride (PVC) products has caused ubiquitous contamination to the environment and potential ecology security risk all around the world, especially in places plastic films were indispensably utilized due to the widely proposing of facility agriculture in China. A case of PAEs contamination in four suburb areas of Nanjing was analyzed and discussed in this study. A new frame work has been put forward based on multi-criteria evaluation model and mathematical method of catastrophe theory, using farming work, laboratory determination and relevant environmental standards to measure the ecology security risk of PAEs in study areas. The factors were selected based on the availability of the data and the local conditions. The assessment model involves the contamination status of PAEs in soil and vegetables, the contamination effects of PAEs to human and soil organisms and the contamination source of PAEs from plastic films and other products in the four study facility agriculture areas. An evaluation system of the model was composed of thirteen mesosphere indicators and twenty-five underlying indicators including total PAEs concentration in soils, single PAE concentration in soils, total PAEs concentrations in roots, leafy, solanaceous and stem vegetables, PAE human risks, soil microbial counts, microorganism diversity indices, atmospheric deposition of PAEs, whether sewage wastewater irrigation, planting mode of the facility agriculture areas and climate condition of study areas. The modified evaluation system was used in the assessment of ecology security of the same place based on the data of 2012, and the results suggested that the ecology security indicators were reliable and were agree well with the practical situation of the study areas. The results could provide guidance for the application of health risk assessment of soil environment for the strong objectivity of catastrophe theory compared with other evaluation methods.

Assessment tools for microplastics and natural fibres ingested by fish in an urbanised estuary.

Microplastics and fibres occur in high concentrations along urban coastlines, but the occurrence of microplastic ingestion by fishes in these areas requires further investigation. Herein, the ingestion of debris (i.e., synthetic and natural fibres and synthetic fragments of various polymer types) by three benthic-foraging fish species Acanthopagrus australis (yellowfin bream), Mugil cephalus (sea mullet) and Gerres subfasciatus (silverbiddy) in Sydney Harbour, Australia has been quantified and chemically speciated by vibrational spectroscopy to identify the polymer type. Ingested debris were quantified using gut content analysis, and identified using attenuated total reflectance Fourier transform infrared (ATR-FTIR) and Raman microspectroscopies in combination with principal component analysis (PCA). The occurrence of debris ingestion at the time of sampling ranged from 21 to 64% for the three species, and the debris number ranged from 0.2 to 4.6 items per fish for the different species, with ∼53% of debris being microplastic. There was a significant difference in the amount of debris ingested among species; however, there was no difference among species when debris counts were standardised to fish weight or gut content weight, indicating that these species ingest a similar concentration of debris relative to their ingestion rate of other material. ATR-FTIR microspectroscopy successfully identified 72% of debris. Raman spectroscopy contributed an additional 1% of successful identification. In addition, PCA was used to non-subjectively classify the ATR-FTIR spectra resulting in the identification of an additional 9% of the debris. The most common microplastics found were polyester (PET), acrylic-polyester blend, and rayon (semi-synthetic) fibres. The potential of using Raman microspectroscopy for debris identification was investigated and provided additional information about the nature of the debris as well as the presence of specific dyes (and hence potential toxicity).

Ingestion of micro- and nanoplastics in Daphnia magna - Quantification of body burdens and assessment of feeding rates and reproduction.

Evidence is increasing that micro- and nanoplastic particles can have adverse effects on aquatic organisms. Exposure studies have so far mainly been qualitative since quantitative measurements of particle ingestion are analytically challenging. The aim of this study was therefore to use a quantitative approach for determining ingestion and egestion of micro- and nanoplastics in Daphnia magna and to analyze the influence of particle size, exposure duration and the presence of food. One week old animals were exposed to 2 µm and 100 nm fluorescent polystyrene beads (1 mg/l) for 24 h, followed by a 24 h egestion period in clean medium. During both phases body burdens of particles were determined by measuring the fluorescence intensity in dissolved tissues. Ingestion and egestion were investigated in the absence and presence of food (6.7·105 cells of Raphidocelis subcapitata per ml). Furthermore, feeding rates of daphnids in response to particle exposure were measured as well as effects on reproduction during a 21 days exposure (at 1 mg/l, 0.5 mg/l and 0.1 mg/l) to investigate potential impairments of physiology. Both particle sizes were readily ingested, but the ingested mass of particles was five times higher for the 2 µm particles than for the 100 nm particles. Complete egestion did not occur within 24 h but generally higher amounts of the 2 µm particles were egested. Animal body burdens of particles were strongly reduced in the presence of food. Daphnid feeding rates decreased by 21% in the presence of 100 nm particles, but no effect on reproduction was found despite high body burdens of particles at the end of 21 days exposure. The lower egestion and decreased feeding rates, caused by the 100 nm particles, could indicate that particles in the nanometer size range are potentially more hazardous to D. magna compared to larger particle sizes.

Assessment of microplastic-sorbed contaminant bioavailability through analysis of biomarker gene expression in larval zebrafish.

Microplastics (MPs) are prevalent in marine ecosystems. Because toxicants (termed here "co-contaminants") can sorb to MPs, there is potential for MPs to alter co-contaminant bioavailability. Our objective was to demonstrate sorption of two co-contaminants with different physicochemistries [phenanthrene (Phe), log10Kow=4.57; and 17α-ethinylestradiol (EE2), log10Kow=3.67] to MPs; and assess whether co-contaminant bioavailability was increased after MP settlement. Bioavailability was indicated by gene expression in larval zebrafish. Both Phe and EE2 sorbed to MPs, which reduced bioavailability by a maximum of 33% and 48% respectively. Sorption occurred, but was not consistent with predictions based on co-contaminant physicochemistry (Phe having higher log10Kow was expected to have higher sorption). Contaminated MPs settled to the bottom of the exposures did not lead to increased bioavailability of Phe or EE2. Phe was 48% more bioavailable than predicted by a linear sorption model, organism-based measurements therefore contribute unique insight into MP co-contaminant bioavailability.

Assessment of models for anaerobic biodegradation of a model bioplastic: Poly(hydroxybutyrate-co-hydroxyvalerate).

Kinetic models of anaerobic digestion (AD) are widely applied to soluble and particulate substrates, but have not been systematically evaluated for bioplastics. Here, five models are evaluated to determine their suitability for modeling of anaerobic biodegradation of the bioplastic poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV): (1) first-order kinetics with and without a lag phase, (2) two-step first-order, (3) Monod (4) Contois, and (5) Gompertz. Three models that couple biomass growth with substrate hydrolysis (Monod, Contois, and Gompertz) gave the best overall fits for the data (R2>0.98), with reasonable estimates of ultimate CH4 production. The particle size limits of these models were then evaluated. Below a particle size of 0.8mm, rates of hydrolysis and acetogenesis exceeded rates of methanogenesis with accumulation of intermediates leading to a temporary inhibition of CH4 production. Based on model fit and simplicity, the Gompertz model is recommended for applications in which particle size is greater than 0.8mm.

Microplastics in the Solent estuarine complex, UK: An initial assessment.

Microplastics are known to be an increasing component found within both marine sediments and the water column. This study carried out an initial assessment of the levels of microplastics present within the Solent estuarine complex, focusing specifically on the water column. A plankton net trawl survey was carried out, with samples analysed using visual observation and Fourier Transform Infrared Spectroscopy (FT-IR). The study identified significant quantities of plastics, ranging in shape, with hot spots found at confluence points within the estuary. Though the FT-IR analysis was inconclusive, the nature of the samples indicates the effect of oceanographic conditions on the prevalent types of microplastics found, which in turn identifies key local sources such as wastewater treatment plants and the plastics industry as being the dominant inputs.

Deterioration of bioplastic carrier bags in the environment and assessment of a new recycling alternative.

Increasing environmental concerns and the introduction of technologies based on renewable resources have stimulated the replacement of persistent petroleum-derived plastics with biodegradable plastics from biopolymers. As a consequence, a variety of products are currently manufactured from bioplastic, including carrier bags. This series of studies investigated the deterioration of carrier bags made with Mater-Bi (MB), a starch-based bioplastic, in soil, compost and two aquatic ecosystems, a littoral marsh and seawater. Results from the laboratory study indicated that bioplastic carrier bags were rapidly deteriorated in soil and compost. After three months of incubation, weight loss of specimens was of 37% and 43% in soil and compost, respectively. Conversely, little deterioration was observed in specimens buried in soil under field conditions or exposed to water of a littoral marsh and of the Adriatic Sea. These findings were consistent with the greater number of bacteria and especially fungi capable of degrading MB that were recovered from soil and compost with respect to the two aquatic ecosystems. Considering that a variety of microbial isolates are capable of using MB as a source of carbon, a new alternative to recycle these MB-based carrier bags was explored. More specifically, starchy residues from bags were fermented by the fungus Rhizopus oryzae to produce up to 35 mg of lactic acid per g of bag residues.

Assessment of morphological and immunohistological alterations in long-term keloid skin explants.

One of the major impediments in keloid research is the lack of a keloid animal model that can mimic human keloid. This imposes investigative constraints on studying cellular interactions and biochemical processes that normally occur in vivo. Our main objective is to establish an in vitro model for maintaining long-term viable keloid dermal explants as a tool for investigating the pathogenesis of keloid scar formation. Explants of adult keloid scars were cultured in vitro by embedding them in enriched collagen gel matrix and maintaining them for up to 6 weeks, whereupon changes in tissue morphology and cellular differentiation were examined. The effects of medium enrichment, air versus liquid submersion, and different substrates on the explants were examined. Our results indicated that keloid explants embedded in a collagen gel matrix were morphologically better preserved than explants placed on a plastic substrate. Explants with epidermis at the air-liquid interface had better morphology than collagen-submerged explants, and there were no differences between serum-free and serum-supplemented explant cultures. Immunohistochemical and apoptotic analyses were performed to assess cellular viability and differentiation. In situ hybridization confirmed that keloid fibroblasts had sustained collagen type I gene expression throughout the 6 weeks in culture, thus validating the integrity of a long-term keloid culture system. In conclusion, the collagen-embedded skin explant system demonstrates that keloid tissues could be maintained for up to 6 weeks for long-term in vitro studies.