Marine antifouling efficacy of amphiphilic poly(coacrylate) grafted PDMSe: effect of graft

molecular weight

Cary A. Kuliashaa, John A. Finlayb, Sofia C. Francob, Anthony S. Clareb, Shane J. Stafslienc, and

Anthony B. Brennana

aDepartment of Materials Science and Engineering, University of Florida, Gainesville, FL, USA;

bSchool of Marine Science and Technology, Newcastle University, Newcastle upon Tyne, UK;

cOffice of Research and Creative Activity, North Dakota State University, Fargo, ND, USA

Supplemental Information

Bulk Copolymer Characterization

GPC analysis was performed on bulk copolymer samples collected from grafting

solutions and purified via dialysis. The Mw of the bulk copolymer was used to estimate the graft

molecular weight as a function of changing the concentration of the chain transfer agent, TGA

(Figure S1). These results indicate that the TGA mediated chain growth polymerization could

produce a wide range of Mw’s; however, the precise control of Mw was lacking as indicated by

high deviations in Mw obtained batch-batch. Better Mw control is possible utilizing living

polymerization techniques; however, it is unlikely that the large range of overall Mw’s produced

utilizing TGA would be possible with living techniques.

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Figure S1. (a) Effect of increasing [TGA] on the MW of bulk copolymer purified from solution

and (b) representative GPC chromatograms of copolymers with different MW values produced by

varying [TGA]. Data points represent the arithmetic mean with error bars representing one

standard deviation, n=4

ATR-FTIR analysis of homopolymers was performed to identify distinguishable peaks

that could be used for copolymer analysis (Figure S2). Copolymer composition was estimated by

performing peak fitting analysis of the carbonyl region (Figure S3) and fitting the respective

heights of each of the three carbonyls normalized by the -CH2 bend to a Beer’s law type relation.

This relation was derived by performing IR analysis of blends of PAA conjugated with Na+

(neutral dialyzed) and PAAm at different ratios (Figure S4). Blends were made by solvating the

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homopolymers in water (pH=7.5), allowing them to mix thoroughly, and drying them to a

homogenous film. The Beer’s law relationships for AA and AAm content show good linear

relationships (Figure S5). PMA was not incorporated into these blends due to its insolubility in

water, and PMA composition was estimated by subtracting the calculated composition of AA

and AAm from 100%. These relationships were validated by performing the same peak fitting

procedure on poly(AA Na+-c-AAm-c-MA) blended with varying concentrations of either PAA or

PAAm.

Figure S2. ATR-FTIR spectra of poly(acrylate) homopolymers. All polymers were synthesized

in lab, purified by dialysis, and dried by rotary evaporation. The 1452 cm-1 peak is the -CH2 bend

attributed to the polymer backbone. Carbonyl designations are as follows: 1732 cm-1 PMA, 1704

cm-1 PAA, 1664 cm-1 PAAm, and 1556 cm-1 PAA Na+.

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Figure S3. ATR-FTIR spectra of a ternary poly(co-acrylate) bulk sample showing the fitted

peaks utilized for copolymer composition analysis.

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Figure S4. ATR-FTIR spectra of a homopolymers, homopolymer blends, and ternary copolymer

utilized to calculate the Beer-Lambert law relationship.

Figure S5. Beer’s law type of relation used for copolymer compositional analysis. The carbonyl

peak height (either 1556 cm-1 for PAA or 1664 cm-1 for PAAm) was normalized with the -CH2

bend at 1452 cm-1.

PCAgPDMS Characterization

The Owens-Wendt method was used to calculate the surface free energy (SFE), γS

(mN/m), of test surfaces using W-MI and MI-GL probe liquid pairs. Equation S1 was used to

calculate the surface’s polar and dispersive (γSp and γS

d) components using the measured contact

angle (θ) of each liquid and polar/dispersive values found in Table S1. The overall SFE, γS, was

determined by averaging the values of the probe liquid pairs obtained using Equation S2.

γ L (1+cos (θ ) )2

=√γ Sd γ L

d+√γ Sp γ L

p (S1)

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γ=γ p+γ d (S2)

Table S1. SFE values for probe liquids used for SFE calculations of PDMSe surfaces.

Probe Liquid γL

(mJ/m2)γL

d

(mJ/m2)γL

p

(mJ/m2)W 72.8 21.8 51.0MI 50.8 48.5 2.3GL 63.4 36.0 27.4

TA.XT Plus texture analyzer by Stable Microsystems using a 5 kg load cell was used to

perform tensile tests on PDMSe test coatings. Testing was performed on dog-bone samples

(soaked in water for >100 h) according to ASTM D412-15A. The elastic modulus (E) was

calculated according to the slope of the stress-strain curve within the elastic regime of the tests.

Grafting caused no change in the elastic modulus compared to PDMSe.

PCAgPDMS coatings adsorbed water due to the hydrophilic nature of the surface grafts

in a fashion not seen for PDMSe coatings, and this swelling resulted in a change in the optical

appearance of the coatings. ATR-FTIR was used to confirm that swelling of water, not some

other unknown reason, caused the change seen in optical appearance and mass. IR analysis of

swollen PCAgPDMS samples was able to detect two intense absorbance bands centered around

3400 cm-1 and 1650 cm-1 consistent with the IR absorbance of H2O (Figure S7). Dehydrating

these samples by either vacuum or desiccation eliminated these water peaks.

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Figure S6. (a) Stress-strain data averaged between three samples per coating, and (b) the same

data restricted to the linear portion within the elastic regime. The Elastic modulus of 1.16 MPa

was consistent for both PDMSe and PCAgPDMS coatings. Both coatings slipped from the

tensile machine’s grips at higher strains as indicated by the large drops in stress seen in part a.

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Figure S7. ATR-FTIR spectra of PCAgPDMS coatings swollen in water (blue) or dessicated

(red) and PDMSe control swollen in water (black). Water adsorption by the PCAgPDMS

samples was due to the acrylate grafts. Inset figures show the two main areas of water

absorbance in the (a) >3000 cm-1 region and (b) the fingerprint region from 1800-1300 cm-1.

PCAgPDMS N. incerta Bioassay Data

Sample leachate toxicity against Navicula incerta was assessed by introducing diatoms

into overnight extracts (ASW with nutrients) of treatment coatings and evaluating growth after

48 h via fluorescence of chlorophyll (Figure S8). Growth in coating leachates was reported as a

fluorescence ratio compared to a positive growth control (fresh nutrient medium) and a negative

growth control (medium + bacteria + 6 μg/ml triclosan). PCAgPDMS samples displayed no

evidence of leachate toxicity; however, IS700 and IS900 samples showed mild toxicity despite

the 7 day tap water immersion. PCAgPDMS coatings did not impact the 48 h biofilm growth

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compared to the PDMSe control, but IS700 and IS900 showed diminished growth likely due to

their mild toxicity.

Figure S8. (a) Coating leachate toxicity towards N. incerta and (b) N. incerta biofilm growth

after 48 h on coatings in 24-well plates. Data is from Bioassay 5, and error bars represent one

standard deviation. The dashed line for part (a) is included as a visual reference for the positive

growth control. Groups that share the same letter are statistically equivalent, α=0.05.

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Figure S9. Initial attachment (2 h settlement) and removal of N. incerta by (a) 138 kPa water jet

impact pressure on coated 24-well plates (bioassay 6), and (b) 26 Pa water shear stress on coated

glass slides (bioassay 8). Inset percentage values represent the percentage removal of diatoms

due to applied water pressure. Error bars represent 95% confidence intervals. Groups that share

the same letter (black or grey) are statistically equivalent, α=0.05.

Table S2. Percentage removal values of N. incerta diatoms performed on 24-well plates between

two bioassays at both impact pressures of 69 and 138 kPa.

Bioassay 5 Bioassay 6

Coating % Removal 69 kPa

% Removal 138 kPa

Coating%

Removal 69 kPa

% Removal 138 kPa

PDMSe 37.3 ± 2.4 59.3 ± 3.9 PDMSe 37.3 ± 4.5 66.3 ± 6.8PCAgPDMS

Mw=994 kg/mol 63.4 ± 8.1 86.4 ± 2.8 PCAgPDMS Mw=1,022 kg/mol 35.9 ± 2.3 74.4 ± 3.1

PCAgPDMS 52.2 ± 8.9 83.5 ± 1.9 PCAgPDMS 49.6 ± 3.9 82.5 ± 4.6

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Mw=835 kg/mol Mw=619 kg/molPCAgPDMS

Mw=441 kg/mol 53.1 ± 7.2 75.6 ± 6.0 PCAgPDMS Mw=397 kg/mol 28.0 ± 7.5 72.0 ± 6.3

PCAgPDMS Mw=221 kg/mol 51.4 ± 4.4 75.3 ± 2.6 PCAgPDMS

Mw=227 kg/mol 43.7 ± 3.8 73.7 ± 1.8

IS700 7.2 ± 2.7 24.5 ± 13 IS700 ----- -----IS 900 23.2 ± 4.9 51.7 ± 2.7 IS 900 9.0 ± 6.6 53.6 ± 10

IS 1100SR 53.0 ± 4.6 88.3 ± 1.2 IS 1100SR 26.3 ± 9.9 77.2 ± 3.0*Percentage removal is reported by comparing each count of diatom biomass remaining post-removal to the mean

diatom initial biomass pre-removal per coating type. Arithmetic average ± 95% confidence interval, n=4. IS 700 was

excluded from bioassay 6 due to issues with coating adherence to well plates.

Table S3. Percentage removal values of N. incerta diatoms performed on coated glass slides for

two bioassays.

Bioassay 7 Bioassay 8

Coating % Removal26 Pa Coating % Removal

26 PaPDMSe 0.0 ± 0.2 PDMSe 10.4 ± 3.9

PCAgPDMS Mw=1,319 kg/mol 39.1 ± 4.2 PCAgPDMS Mw=1,022 kg/mol 34.3 ± 6.1PCAgPDMS Mw=1,190 kg/mol 54.6 ± 3.4 PCAgPDMS Mw=619 kg/mol 35.4 ± 5.4PCAgPDMS Mw=138 kg/mol 46.4 ± 4.9 PCAgPDMS Mw=397 kg/mol 26.7 ± 5.4PCAgPDMS Mw=80 kg/mol 40.9 ± 5.4 PCAgPDMS Mw=227 kg/mol 33.0 ± 6.5

*Percentage removal is reported by comparing each count of remaining diatom density post-removal from 30 counts

on each replicate to the mean initial diatom density pre-removal per coating type, and an arc-sine transformation was

performed to obtain more representative error bars due to the nature of the percentage values. Arithmetic average ±

95% confidence interval, n=3

Statistical Analysis

Parametric one-way ANOVA makes several assumptions such as homogeneity of

variance and normality that were tested for, and the results of these tests as well as the F-

statistics for the ANOVA’s run for all bioassay data is shown in Table S4. The Shapiro-Wilk

test and corresponding Q-Q plots were used to confirm that the data used was normally

distributed. This test was performed on all U. linza attachment density data per coating per

bioassay and all N. incerta initial and remaining biomass data per coating per bioassay. The

results of each test are not reported due to the shear size of the data set (60 individual tests and

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plots). Levene’s test was used tests the null hypothesis that the population variances are equal, to

confirm homogeneity of variance (Table S4). If the null hypothesis was rejected by a p values <

0.05, both the Welch and the Brown-Forsycthe tests were used to determine significance instead

of the ANOVA F-statistic.

Table S4. Statistical Summary

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Bioassay # Data Set Levene’s One-Way

ANOVA Welch Brown-Forscythe

1 Attachment Density F(4,445)=27.282, p=0.000 --------- F(4,216)=228.925,

p<0.05F(4,205)=523.550,

p<0.05

2 Attachment Density F(4,445)=99.039, p=0.000 --------- F(4,218)=238.706,

p<0.05F(4,114)=850.025,

p<0.05

5 Initial Attachment F(7,16)=1.403, p=0.271

F(7,16)=19.425, p<0.05 --------- ---------

5 Remaining Attachment, 69 kPa

F(7,16)=2.128, p=0.100

F(7,16)=51.303, p<0.05 --------- ---------

5 Remaining Attachment, 138 kPa

F(7,16)=2.336, p=0.076

F(7,16)=48.767, p<0.05 --------- ---------

6 Initial Attachment F(6,14)=1.902, p=0.151

F(6,14)=24.495, p<0.05 --------- ---------

6 Remaining Attachment, 69 kPa

F(6,14)=1.185, p=0.369

F(6,14)=13.868, p<0.05 --------- ---------

6 Remaining Attachment, 138 kPa

F(6,14)=1.386, p=0.287

F(6,14)=3.439, p<0.05 --------- ---------

7 Initial Attachment F(4,445)=1.050, p=0.381

F(4,445)=5.581, p<0.05 --------- ---------

7 Remaining Attachment

F(4, 445)=4.827, p=0.001 --------- F(4,220)=78.444,

p<0.05F(4,398)=85.566,

p<0.05

8 Initial Attachment F(4,445)=0.842, p=0.499

F(4,445)=2.965, p<0.05 --------- ---------

8 Remaining Attachment

F(4,445)=3.672, p=0.006 --------- F(4,222)=11.409,

p<0.05F(4,404)=16.319,

p<0.05