Microfibers from Laundering and Their Fate in the Aquatic Environment

Marielis Zambrano1, Richard Venditti1, Joel Pawlak1, Jesse Daystar2, Mary Ankeny2, Carlos Goller3, Jay Cheng4

1Department of Forest Biomaterials, College of Natural Resources, North Carolina State University; 2Cotton Incorporated; 3Department of Biological Sciences, North Carolina State University; 4Department of Biological and Agricultural Engineering, North Carolina State University.

December 3, 2019 OUTLINE

Outline

 Introduction  Microfibers generated from laundering  Aerobic biodegradation of textile spun yarns in aquatic and marine environments.  Microfibers interaction with the microbiome  Summary

www.adventurescientists.org/microplastics.html

2 MOTIVATION

Microplastics: Sources and Distribution

Microplastics are EVERYWHERE!!! “Microplastics are any synthetic solid particle or polymeric matrix, with regular or irregular shape and with Beer Tap Water Sea Salt size ranging from 1 μm to 5 mm, of either primary or secondary manufacturing origin, which are insoluble in water”. Frias and Nash (2019)

About 0.8 - 2.5 Mt./year of primary microplastics are 212 particles/kg released into the ocean 4 particles/L 5 particles/L The average person ingests over 5,800 particles of synthetic 35% are micro-size fibers released from textiles during debris from these three sources annually, laundering. Kosuth et al. (2018) The presence of anthropogenic debris in seafood for human consumption has been observed

Miranda and de Carvalho-Souza (2016) and Rochman et al. (2015) Human stool samples have shown between 18 to 172 plastic particles per 10 g of stool.

Schwabl (2018)

Boucher and Friot (2017) 3 Textile Microfibers in the Environment

Wastewater Treatment Plants The Lint LUV-R filter traps 87% of fibers

Washing Machine Effluent

High volumes discharged daily Small size (10 µm – 100 µm) 30% Fibers

WWTP Home Laundering WWTP Effluents The Coral Ball traps 26% of fibers > 98 % McIlwraith et al. (2019) Microplastics (MPs) Removal Efficiency

Magnusson et al. (2014), Lares et al. (2018), Talvitie et al. (2015), Talvitie et al. (2017a) Talvitie et al. (2017b), Mason et al. (2016), Mintenig et al. (2017), Wolff et al. (2018). Environmental Impact Textiles Rummel et al. (2017) 4 OBJECTIVES Objectives

Quantify microfibers generated from laundering of different fabrics

Understand biodegradation of microfibers in aquatic environments.

Understand the impact of microfibers on the microbial communities of lake and seawater ecosystems.

5 Microfibers generated from the laundering of cotton, rayon and polyester based fabrics

Do synthetic fabrics release a different amount of microfibers during laundering than natural fabrics?

What are the washing parameters that influence the most the microfiber release?

What is the mechanism of microfiber release during laundering? METHODOLOGY

Accelerated Laundering Experiments

Quantify

SDL Atlas Launder-Ometer

Fibers Quality http://www.sdlatlas.com/product/120/La Water with http://www.sdlatlas.com/consumable/81/Container- under-Ometer Seals-(pack-of-8) Microfibers Analyzer HiRes FQA Washing Cycle (Robertson et al. 1999) Samples Containers Preparation Laundering Water Collection Preparation 16 min • 1 piece of fabric / Container At constant temperature • 25 metal balls / Container Weft Knitted Interlock . • 150 ml Detergent Solution Fabric 8 Containers / Washing Cycle (1.47g AATCC Standard Filtration and Liquid Detergent per 1L of 4 in x 4 in Weighing Pre-Cleaned DI Water) or DI water

7 Robertson, G., Olson, J., Allen, P., Chan, B. E. N., and Seth, R. (1999). “Measurement of fiber length, coarseness, and shape with the fiber quality analyzer,” Tappi Journal, 82(10), 93–98. RESULTS Accelerated Laundering Experiments High Temperature Low Temperature No detergent Detergent No detergent Detergent

Cotton Polyester 50/50 Cotton/Polyester Rayon

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 8 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 RESULTS

Microfibers Size Distribution Fibers Length= 0.2mm - 10mm  Microfibers shed from laundering detected in our 16% experiments .02-2 mm and about .016 mm wide. Cotton = Purple 12% Rayon = Green Polyester = Pink Polyester/Cotton = Orange  Fibers used to spin yarns > 25 mm. 8%

 This indicates that the textile fibers are broken during 4%

washing. Normalized Distribution 0%  Small size: easy to ingest……hard to detect. 0.00 - 0.05 0.15 - 0.20 0.30 - 0.35 0.45 - 0.50 0.60 - 0.65 0.75 - 0.80 0.90 - 0.95 1.05 - 1.10 1.20 - 1.25 1.35 - 1.40 1.50 - 1.55 1.65 - 1.70 1.80 - 1.85 1.95 - 2.00 2.10 - 2.15 2.25 - 2.30 2.40 - 2.45 2.55 - 2.60 2.70 - 2.75 Length (mm)

Cotton 50/50 Polyester/Cotton Polyester Rayon

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 9 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 METHODOLOGY

Microfibers Generated During Home Laundering

4 Lb of Fabric Cycles Pre-Cleaned 700,000 microfibers released per Wet Fabric load Washing Drying Dried Fabric

Normal Cycle High (Heavy) Cycle 60 min

45 gallons Container

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 10 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 RESULTS

Microfibers are broken from the yarn during washing

100 % Polyester 50/50 Polyester/Cotton Spun Yarn Spun Yarn

100 % Rayon 100 % Cotton Spun Yarn Spun Yarn

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 11 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 RESULTS

Microfibers are broken from the yarn during washing

100 % Polyester 50/50 Polyester/Cotton Spun Yarn Spun Yarn

100 % Rayon 100 % Cotton Spun Yarn Spun Yarn

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 12 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 Aerobic Biodegradation of Textile Spun Yarns in Aquatic Environments

Do microfibers released during laundering biodegrade in aquatic and marine environments? What is biodegradation?

 A chemical compound is transformed or eliminated by the biological action of living organisms.

(Science Photo Library, fineartamerica)

14 What are the steps for biodegradation?

1. For bacteria to live they need food (substrate), pH, temperature, oxygen, water 2. Bacteria must come into proximity of Food Source (microfibers) the substrate 3. The bacteria secretes enzymes that adsorb onto the substrate and catalyze the breakage of compounds into small units such as sugars 4. The sugars diffuse to the bacteria and are transported through the cell wall 5. Sugars are processed by the bacteria, consuming oxygen, producing biomass, energy and carbon dioxide Bacteria

15 METHODOLOGY Aerobic Biodegradation of Textile Spun Yarns in Aquatic and Marine Environments

ISO 14851:1999. Determination of the ultimate aerobic biodegradability of plastic materials in an aqueous medium -- Method by measuring the oxygen demand in a closed respirometer; 2005. 16 ASTM D6691 − 09. Standard Test Method for Determining Aerobic Biodegradation of Plastic Materials in the Marine Environment by a Defined Microbial Consortium; West Conshohocken, PA, 2009. RESULTS

Aerobic Biodegradation of Textile Spun Yarns (Lake and Seawater)

95 100% Cotton 100% Polyester 95 100% Cotton 100% Polyester 85 100% Rayon 50/50 Polyester/Cotton 85 100% Rayon 50/50 Polyester/Cotton MCC 75 75 MCC 65 65 55 Rayon Cotton 55 Cotton 45 45 35 35 Rayon 25 25 50/50 % Biodegradation % 50/50 Biodegradation % 15 15 5 Polyester 5 -5 -5 Polyester 0 5 10 15 20 25 30 35 0 5 10 15 20 25 30 Days Days Lake Water Seawater

ISO 14851 ASTM D6691 Determination of the Ultimate Aerobic Biodegradability of Plastic Materials in an Aqueous Determination of the Ultimate Aerobic Biodegradability of Plastic Materials in an Medium Aqueous Medium N=3 N=4 Inoculum – Lake Raleigh Water Inoculum – Seawater Measurements – RSA PF-8000 (Oxygen Uptake) Measurements – RSA PF-8000 (Oxygen Uptake) Material Added – 80 mg of yarns/500 ml Test Medium Material Added – 80 mg of yarns/500 ml Test Medium

Zambrano, M. C.; Venditti, R. A.; Pawlak, J. J.; Ankeny, M.; Daystar, J.; Goller, C. Aquatic Biodegradability of Cotton, Polyester, and Rayon Yarns (2nd Place Winner, Herman and Myrtle Goldstein Graduate Student Paper Competition). 17 AATCC 2019 International Conference (April 2019). Texas, United States. Available online: https://www.aatcc.org/wp-content/uploads/2019/03/Zambrano-Marielis-HMGSPC.pdf RESULTS

Aerobic Biodegradation of Textile Spun Yarns SEM images of the spun yarns during biodegradation using as inoculum 30 ppm of Activated Sludge solids from the Neuse River WWTP 100% Cotton 100% Polyester 100% Rayon 50/50 Polyester/Cotton Day 0 Day Day 6 Day Day 38 Day 18 OBJECTIVES

Effect of Finishes on Cotton Aquatic Biodegradation

Cellulose Study the effect of typical textile finishes (dye, durable press, softener, and water repellent) on the biodegradation of cotton in aquatic environments.

Cotton – Dyed Cotton – Durable Press Cotton – Softener Cotton – Water Repellent Reactive Blue 19 DMDHEU & Catalyst Modified amino functional C6 & PBI silicone Non-PFOA fluorochemical Polyfunctional blocked isocyanate crosslinker

19 RESULTS

Effect of Finishes on Cotton Biodegradation in Aquatic Environments

110 100 90 80 70 60 50 40 Blue 19 30 Cotton - No Finish Cotton - Softener

% Biodegradation % 20 Cotton - Dyed (Blue 19) Cotton - Durable Press 10 Reference Material (MCC) Cotton - Water Repellent 0 Oak Leaves 0 20 40 60 80 100 Days 30 ppm of Activated Sludge solids from the Neuse River WWTP ISO 14851 Determination of the Ultimate Aerobic Biodegradability of Plastic Materials in an Aqueous Medium Durable Press N=3 all materials, except Oak Leaves (N=2) Measurements – RSA PF-8000 (Oxygen Uptake) Material Added – 100 mg of yarns/500 ml Test Medium 20 ISO 14851:1999. Determination of the ultimate aerobic biodegradability of plastic materials in an aqueous medium -- Method by measuring the oxygen demand in a closed respirometer; 2005. How do Textile Microfibers Interact with the Microbiome in Aquatic Environments?

Microbiome – the community of microorganisms that inhabit an environment and/or the collective genomes of the microorganisms

The microbiome of the human gut provides human metabolism, nutrition, physiology, and immune function.

Source: Definition of Microbiome. https://www.merriam- webster.com/dictionary/microbiome [accessed 3/8/2018] A human has about 2.8 x 1013 bacteria cells. Integr Med (Encinitas). 2014 Dec; 13(6): 17–22. About the same as the number of human cells.

Source: Sender R, Fuchs S, Milo R. (2016). Revised Estimates for the Number of Human and Bacteria Cells in the Body. August 19, 2016. PLOS. https://doi.org/10.1371/journal.pbio.1002533 Source: Definition of microbiota. https://www.merriam-webster.com/dictionary/microbiota [accessed 3/8/2018] METHODOLOGY

Microbiome Analysis

MultiQC NEPHELE DADA2 Amplify DNA from 16S rRNA regions Sequence using Isolate/Purify from Genomic Sample Collection Data Analysis Genomic DNA DNA and barcode Next Generation samples with Sequencing Illumina indexes. Illumina Phyloseq Bacteria MiSeq 300 PE DeSeq2 Origin

22 RESULTS Differential Abundance Analysis Relative to the Blank Top 20 Most Abundant Bacterial Groups

Lake Water Biodegradation Experiment Seawater Biodegradation Experiment

23 RESULTS Beta Diversity Principal coordinate analysis (PCoA) of 16S rRNA gene libraries based on UNIFRAC Method (weighted)

Initial Initial Inoculum Inoculum

Biodegradable Samples Biodegradable (Cotton) Samples (Cotton)

Non- Non- Degraded Degraded Samples Samples (Polyester) (Polyester)

Lake Water Biodegradation Experiment Seawater Biodegradation Experiment

Cellulose based materials (cotton) were a food source for naturally occurring micro-organisms and promoted their abundance.

Non-degradable materials (polyester) did not interact with the micro-organisms. 24 Concluding Remarks

 Laundering of fabrics is a significant contributor to the microfiber issue. (700,000 microfibers/load)

 All fabric fiber types tested shed microfibers (cotton, rayon, polyester)

 Cellulose-based cotton biodegrades quickly in aerobic aquatic environments and polyester does not.

 Cotton and polyester based microfibers promote DIFFERENT changes in the microbiome.

 Quantity of microfibers and their environmental persistence/impact both need to be considered with respect to the environment:  Both should be considered in test methods and decisions. 25 Acknowledgements

Department of Forest Biomaterials

Environmental Analysis Lab NC State University Genomic Sciences Laboratory

Thank you!

26 RESULTS

Aerobic Biodegradation of Textile Spun Yarns (WWTP Solids)

100% Cotton 50%/50% Polyester/Cotton 120 100% Polyester 100% Rayon Reference Material (MCC) 100 Cotton 80 Rayon

60 50/50 Cotton/Polyester

% Biodegradation % 40

0 Polyester 0 5 10 15 20 25 30 35 40

Days

30 ppm of Activated Sludge solids from the Neuse River WWTP ISO 14851 Determination of the Ultimate Aerobic Biodegradability of Plastic Materials in an Aqueous Medium N=4 Measurements – RSA PF-8000 (Oxygen Uptake) Material Added – 80 mg of yarns/500 ml Test Medium 27 RESULTS

Microfibers Size Distribution

Cotton Polyester

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 28 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 RESULTS Differential Abundance Analysis Relative to the Blank Top 20 Most Abundant Bacterial Groups

Lake Water Biodegradation Experiment Seawater Biodegradation Experiment

29 RESULTS

Accelerated Laundering vs Home Laundering

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 30 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 RESULTS

Predictive Tests for the Microfibers Generation during Laundering

ASTM D4966 Abrasion Resistance of Textiles Fabrics ASTM D2256 Tensile Properties of Yarns by the Single-Strand Method James Heal Maxi-Martindale Abrasion Tester Tensile Tester MTS Q Test 5 Weight Loss of the fabrics after 20000 cycles in the Martindale Abrasion Tester Breaking Load of the spun yarns after 5 min inversion in DI water against standard wool abrasion fabric

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 31 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 RESULTS Accelerated Laundering Experiments Effect of Detergent Use

44 °C

25 °C

Zambrano, M.C., Pawlak, J.J., Daystar, J., Ankeny, M., Cheng, J.J., Venditti, R.A., 2019. Microfibers generated from the laundering of cotton, rayon and polyester based fabrics and their aquatic biodegradation. Mar. Pollut. 32 Bull. 142, 394–407. https://doi.org/10.1016/J.MARPOLBUL.2019.02.062 RESULTS

Aerobic Biodegradation of Textile Spun Yarns (Lake and Seawater)

95 100% Cotton 100% Polyester 95 100% Cotton 100% Polyester 85 100% Rayon 50/50 Polyester/Cotton 85 100% Rayon 50/50 Polyester/Cotton MCC 75 75 MCC 65 65 55 Cotton 55 Cotton 45 45 35 35 25 25 % Biodegradation % 15 Biodegradation % 15 5 Polyester 5 -5 -5 Polyester 0 5 10 15 20 25 30 35 0 5 10 15 20 25 30 Days Days

Lake Water Seawater

Zambrano, M. C.; Venditti, R. A.; Pawlak, J. J.; Ankeny, M.; Daystar, J.; Goller, C. Aquatic Biodegradability of Cotton, Polyester, and Rayon Yarns (2nd Place Winner, Herman and Myrtle Goldstein Graduate Student Paper Competition). 33 AATCC 2019 International Conference (April 2019). Texas, United States. Available online: https://www.aatcc.org/wp-content/uploads/2019/03/Zambrano-Marielis-HMGSPC.pdf