Antibacterial Property and Biodegradation of PLA/PBS Nonwoven Fabric Coated with Mangosteen Pericarp Extract

In light of the eco-friendliness and antimicrobial property of mangosteen pericarp (MP), this research investigates the antibacterial activity of biodegradable poly(lactic acid) (PLA)/poly(butylene succinate) (PBS) nonwoven fabric coated with MP extract. In the experiment, the MP extract concentrations were varied between 0 (uncoated), 0.3, 0.5, 1, 3, and 5 wt%, and the experimental bacteria strains were Escherichia coli and Staphylococcus aureus. The results revealed that the MP extract inhibited the growth of E. coli and S. aureus. The tensile strength and the MP extract concentration were positively correlated, while the elongation at break was inversely correlated with the MP extract concentration. Meanwhile, higher MP extract concentrations had minimal effect on the biodegradability of the MP-coated PLA/PBS nonwoven fabrics. Essentially, the MP extract holds promising potential as an antibacterial. The characteristic of this research lies in the use of mangosteen pericarp extract, an agricultural waste, in biodegradable nonwoven fabric to inhibit the bacterial growth.


Introduction
Non-biodegradable nonwoven fabrics have been used in a wide range of products because of affordability, versatility, availability, disposability, and low lint [1]. In medical services, nonwoven fabrics are used to reduce the risk of cross infection and maintain high standards of hygiene. There has been growing interest in the study of the functional properties of nonwoven fabrics, especially the antimicrobial properties [2]. antiulcerogenic, antifungal, antibacterial, anticancer, antioxidant, and anti-inflammatory properties [13][14][15].
With growing concern over global warming and environmental degradation, biodegradable polymers, such as poly(lactic acid) (PLA) and poly(butylene succinate) (PBS), are increasingly adopted as sustainable alternatives to conventional non-biodegradable polymers [16]. The blending of PBS with PLA enhances the toughness of fibers while retaining the biodegradable property [17]. PBS also possesses high flexibility, high impact strength, and high thermal and chemical resistance [18,19]. However, PLA/PBS blend is immiscible due to phase separation caused by crystallization and low interfacial adhesion. To overcome the immiscibility, the ratio of PLA to PBS of this current research was thus 80:20 wt% [20,21]. Recently, antibacterial compounds have been added into products made from biodegradable polymers to study the antibacterial activity. Toncheva, A., et al. studied the antibacterial activity of electrospun mats based on polylactide (PLA) against gram-positive (S. aureus) and gram-negative (E. coli) bacteria. 5-nitro-8-hydroxyquinoline (5N8Q) was loaded as an antibacterial drug. It was found that the addition of 10 wt% 5N8Q in PLA fibers showed antibacterial activity and well-defined inhibition zones [22]. Hongsriphan, N., et al. studied antibacterial food packaging sheets from biodegradable PLA/PBS blend coated with chitosan. It was found that the antibacterial activity against E. coli and S. aureus were increased upon the increasing in chitosan concentration [23]. Sukthavorn, K., et a.l studied the antibacterial activity of PLA blended with ground tea leaves (GTL) as a filler. It was found that the PLA composite fabric added GTL could inhibit the bacteria S. aureus, B. subtilis, and C. albicans at up to 65.57, 94.44, and 7.69%, respectively [24].
This research was focus on 2 main areas comprises of (1) study on the antibacterial property of mangosteen pericarp extract coated on PLA/PBS nonwoven fabric, and (2) the production process of nonwoven from PLA/PBS blends and the biodegradability of the nonwoven fabric. The effect of mangosteen pericarp extract on biodegradation rates of PLA/PBS nonwoven fabric was studied. The laboratoryscale disintegration tests of uncoated and MP-coated PLA/ PBS nonwoven fabrics were carried out under composting condition in accordance with the ISO 20200 standard.
Specifically, this research comparatively investigates the antibacterial activity of biodegradable PLA/PBS nonwoven fabric coated with MP extract of variable concentrations: 0 (uncoated), 0.3, 0.5, 1, 3, and 5 wt%. The experimental bacteria strains were Escherichia coli and Staphylococcus aureus. Fourier transform infrared spectrometry was used to analyze the chemical composition of the uncoated and MPcoated PLA/PBS nonwoven fabrics. Field emission scanning electron microscopy and thermogravimetric analysis were used to determine the morphology and thermal stability of the experimental nonwoven fabrics. This research also investigated the biodegradation of the uncoated and MPcoated PLA/PBS nonwoven fabrics.

PLA/PBS Blend and Nonwoven Fabric
In this research, poly(lactic acid) (PLA) pellets are Ingeo ™ biopolymer 6100D with a melt flow index of 24 g/10 min (210 °C) and a density of 1.24 g/cm 3 (NatureWorks, LLC). Poly(butylene succinate) (PBS) pellets are BioPBS™ FZ 71PM with a melt flow index of 22 g/10 min (190 °C) and a density of 1.26 g/cm 3 (PTT MCC Biochem Co., Ltd.). PLA and PBS were oven-dried for 12 h at 80 °C prior to compounding in a twin screw extruder (Type 16.55-2N440010, Brabender). The barrel temperatures were 130-190 °C with a screw speed of 30 rpm. The ratio of PLA to PBS in the PLA/PBS blend was 80:20 wt% [20,21].
The PLA/PBS blend was oven-dried for another 12 h at 80 °C and fabricated using a melt jet spinning machine outfitted with a die of three 0.4 mm spinnerets and a hot air outlet at the center (SR-Ruder Bambi SRV-P70/62; Nihon Yuki Co. Ltd.,). The die temperature was 280 °C with an air pressure of 0.5 MPa. The screw speed was maintained at 10 rpm with a constant hot air temperature of 600 °C. The collector rotated at 0.5 m/min with the left-right motion of 25 rpm. The distance between the die and the collector was 60 cm.
The extraction of mangosteen pericarp (MP) was carried out using an ethanol-water solution (40:60) at room temperature for 48 h and oven-dried at 40 °C for 12 h to obtain the MP extract powder. The MP extract powder was dissolved in absolute ethanol prior to spray-coating. The PLA/PBS nonwoven fabric was spray-coated with MP extract (The 2.5 ml/cm 2 volume per unit area) and oven-dried at 40 °C for 2 h. The MP extract concentrations (i.e., ethanolic MP extract) were varied between 0.3, 0.5, 1, 3, and 5 wt%. The chemical structure of α-, β-, and γ-Mangostins in xanthones derived from mangosteen pericarp are shown in Figure 1.

Analytical Method
The chemical functional groups were determined using Fourier transform infrared (FTIR) spectrometry (Thermo Scientific, Nicolet 6700) between the wavenumber of 4,000-500 cm − 1 . The fiber size was measured using field emission scanning electron microscopy (FE-SEM; JEOL, JSM − 7610FPlus, Japan) with an accelerated voltage of 15 kV. The uncoated (0 wt%) and MP-coated PLA/PBS nonwoven fabric specimens were gold-sputtered prior to the FE-SEM analysis, and the ImageJ imaging processing program was used to measure the fiber diameter and size distribution. The thermal stability of the uncoated and MPcoated PLA/PBS nonwoven fabrics was determined using a thermogravimetric analyzer (TGA, Q50), whereby 10 mg of the nonwoven fabrics were heated from 30 to 600 °C at a rate of 10 °C/min under nitrogen gas to prevent oxidation.

Tensile test
The tensile properties of uncoated and MP-coated PLA/ PBS nonwoven fabrics (i.e., tensile strength and elongation at break) were determined by a universal testing machine (Instron, model 5569, USA) at room temperature. The nonwoven fabric specimens were 1 × 6 cm (W × L) in dimension. The grip distance was 25 mm with a testing speed of 50 mm/min in accordance with the ASTM D3822-01 standard.

Antibacterial test
The antibacterial test was carried out qualitatively using agar disc diffusion method to evaluate the antibacterial activity of MP extract coated on the PLA/PBS nonwoven fabric. The experimental bacterial strains were E. coli (Gram-negative, TSM11) and S. aureus (Gram-positive, TSM06). The bacteria were initially incubated in a shaker containing tryptic soy broth at 37 °C for 24 h. Afterward, the bacteria (10 8 CFU/mL) were transferred to Petri dishes containing 15 mL tryptic soy agar (TSA; 30g TSA/1 L water) by swabbing all over the TSA surface.
The uncoated (control) and MP-coated PLA/PBS nonwoven fabric specimens were of circular shape and 25 mm in diameter. The fabric specimens were placed on the top of TSA containing the bacteria and incubated at 37 °C for 24 h. The contact area where the fabric specimens contacted the TSA was visually inspected to determine the antibacterial activity of the uncoated and MP-coated PLA/PBS nonwoven fabrics.

Disintegration test Under Composting Condition
The laboratory-scale disintegration tests of uncoated and MP-coated PLA/PBS nonwoven fabrics were carried out under composting condition in accordance with the ISO 20200 standard. The PLA/PBS nonwoven fabric specimens (25 mm × 25 mm) were buried at 4-6 cm depth in perforated plastic containers containing solid synthetic wet waste and aerobically incubated at 58 °C for 1, 4, 10, 16, 23, and 28 days. The solid synthetic wet waste consisted of 10% compost, 30% rabbit food, 10% starch, 5% sugar, 1% urea, 4% corn oil and 40% sawdust and approximately 55 wt% of water content.
The physical disintegration of PLA/PBS nonwoven fabric specimens were determined at days 1, 4, 10, 16, 23, and 28 and results compared. In addition, the degree of disintegration (D) of the uncoated and MP-coated PLA/PBS nonwoven fabrics was calculated by Eq. (1).
where m i is the initial dry mass (weight) at day 0 and m r is the dry mass (weight) at days 1, 4, 10, 16, 23, and 28.  PLA/PBS fabrics. In comparison, the residual weight (%) of the MP-coated PLA/PBS nonwoven fabrics (0.3, 0.5, 1, 3, 5 wt%) were higher than that of the uncoated nonwoven fabric (0 wt%; control). Besides, the residual weight was positively correlated with the MP extract concentrations. The finding could be attributed to the phenolic compounds in the mangosteen pericarp (e.g., tannins, flavonoids, xanthones). These phenolic compounds have aromatic rings in the backbone, resulting in the rigid structure with high mechanical strength and thermal stability [28]. Figure 4 shows the FE-SEM images of uncoated (0 wt%) and MP-coated (0.3, 0.5, 1, 3, 5 wt%) PLA/PBS nonwoven fabrics. Given 0.3-1 wt% MP extract, the fiber surface appeared smooth, as evidenced by the absence of tiny droplets on the fibers. The smooth fiber surface could be attributed to low viscosity of the MP coating solution. Meanwhile, the tiny droplets appeared on the fiber surface as the MP extract concentrations increased to 3-5 wt%, which due to the ethanol evaporation from the high viscosity solution [29].  Due to 1 H-NMR and FTIR spectra of the extracted MP have been added to the supporting information of this work as the attached file. From the 1 H-NMR spectra as displayed in Fig. S3 -S5, it was found that the extracted MP used in this work consists of the aromatic rings shows the characteristic peaks corresponding to the chemical structures of 1,3,6-trihydroxy-7-methoxy-2,8-diprenylxanthone (α-Mangostin), 1,6-dihydroxy-3,7-dimethoxy-2,8-diprenylxanthone (β-Mangostin), and 1,3,6,7-tetrahydroxy-2,8-diprenylxanthone (γ -Mangostin), respectively. In addition, the FT-IR spectrum of the extracted MP as shown in Fig. S6. It was found that the extracted MP shows a peak at 2964 cm − 1 corresponding to C-H in an aromatic ring, and the peaks at 1455.7 and 1452 cm − 1 correspond to C = C in an aromatic ring on the MP structure. Table 1 tabulates the thermal degradation temperatures at 10%, 50% and 80% of uncoated and MP-coated nonwoven  The experimental results showed the growth of the bacteria around the uncoated (0 wt%) and MP-coated nonwoven fabrics (0.3, 0.5, 1, 3, 5 wt%), indicating the absence of diffusible action. The uncoated PLA/PBS nonwoven fabric (i.e., underneath the nonwoven fabric) exhibited no antibacterial activity, while the antibacterial activity of the MP-coated nonwoven fabrics varied, depending on the MP extract concentrations. In comparison, the MP extract was more effective against S. aureus than E. coli. The phenomenon could be attributed to the double membrane structure of E. coli, which was more complex than the single membrane structure of S. aureus [31]. Koh, J.J. et al. reported that the in vitro antimicrobial activity of xanthones isolated increase in the tensile strength of the coated fabric, compared to the uncoated one. It is noted that the tensile strength of coated fabrics is significantly increased when the MP concentration is coated more than 0.5 wt% would adhere more to the fibers. The coated fabrics at 1 and 3 wt% MP show the highest tensile strength. The increase in the tensile strength of MP-coated PLA/PBS nonwoven fabric might be explained by a good adhesion between hydroxyl groups of MP particle interacted with polar species (C = O) of chemical structure of PLA and PBS in the nonwoven fabrics surfaces that promoted MP adhesion on the PLA/PBS nonwoven fabrics surfaces. However, the tensile strength of the coated fabric is insignificant dropped after 5 wt%. The finding could be attributed to poor adhesion of MP extract  could be attributed to the haphazard structure of PLA/PBS nonwoven fabric (Figure 4). The haphazard and loosely packed structure limits the contact areas between the MPcoated nonwoven fabric and the bacteria [35]. In this research, the disintegration tests of the uncoated and MP-coated PLA/PBS nonwoven fabrics were carried out under composting condition in accordance with the ISO 20200 standard. Figure 7 depicts the disintegration of the uncoated (0 wt%) and MP-coated (0.3, 0.5, 1, 3, 5 wt%) PLA/PBS nonwoven fabrics at days 0, 1, 4, 10, 16, 23, and 28. The disintegration was clearly visible after 10 days under composting condition. Figure 8 shows the degree of disintegration of uncoated and MP-coated PLA/PBS nonwoven fabrics under composting condition. The uncoated PLA/PBS nonwoven fabric (0 wt%) was decomposed by 93% after 28 days. In comparison, the disintegration of the MP-coated PLA/PBS nonwoven fabrics were noticeably slower, depending on the MP from mangosteen peel showed α-mangostin was the most potent plant extract after screening for gram-positive bacteria. The α-Mangostin showed effective effect on destroying the cell walls of S. aureus causing the cells to die [32]. The 1,4-benopyrone structure of α-mangostin was responsible for restricting the bacteria. It was suggested that the mode of action for inhibiting the bacteria follows a three-step process. First, the cytoplasmic membrane is disrupted, causing increased permeability. This is followed by the restriction of the β-lactamase activity and, lastly, the impairment of the peptidoglycan [33]. α-Mangostin has effect on destroying the cell walls of S. aureus causing the cells to die [32]. Because the higher susceptibility of gram-positive bacteria could be related to differences in the cell wall structure, cell physiology, metabolism or degree of contact [34].

Results and Discussion
The antibacterial activity of the 3-5 wt% MP-coated PLA/PBS nonwoven fabrics resembled that of the nonwoven fabrics coated with 0.3-1 wt% MP extract. The finding

Conclusion
This research investigated the antibacterial activity of biodegradable PLA/PBS nonwoven fabric coated with MP extract against E. coli and S. aureus. The MP extract concentrations were varied between 0 (uncoated), 0.3, 0.5, 1, 3, and 5 wt%. The results showed that the uncoated PLA/ PBS nonwoven fabric (0 wt%) exhibited no antibacterial activity, while the antibacterial activity of the MP-coated nonwoven fabrics varied, depending on the MP extract concentrations. The MP extract was more effective against S. aureus than E. coli due to the double membrane structure of E. coli which is more complex than the single membrane structure of S. aureus. The tensile strength and the MP extract concentration were positively correlated, while the elongation at break was inversely correlated with the MP extract concentration. Furthermore, the disintegration of the MP-coated PLA/PBS nonwoven fabrics were noticeably slower, in comparison with the uncoated nonwoven fabric. The delayed disintegration of the MP-coated PLA/ PBS nonwoven fabrics could be attributed to the nonpolar compounds in xanthones coated on those fabrics preventing water diffusion through the polymer matrix. Furthermore, aromatic rings and side groups in xanthones might be resistant to enzymatic hydrolysis in aerobic bacterial degradation. Essentially, the MP extract holds promising potential as an antibacterial. extract concentrations. The delayed disintegration of the MP-coated PLA/PBS nonwoven fabrics could be attributed to the nonpolar compounds such as α-, β-, and γ-Mangostins in xanthones derived from MP [36]. The degradation of polyester is determined not only by the properties of the specimen (crystallinity, molecular weight, chemical structure, and so on) but also by the presence of water or moisture, which is crucial for the growth of microorganisms in the degradation environment and an increasing the rate of hydrolytic degradation [37]. The coating xanthones on those fabrics hindered the diffusivity of water inside the matrix due to its hydrophobic nature leading to the delayed degradation of those MP-coated fabrics. In addition, the presence of aromatic rings and side groups in xanthones might provide a sterical hindrance for an enzymatic attack to those fabrics resulting in resistance to enzymatic hydrolysis in aerobic bacterial degradation. Furthermore, carbonyl group content in xanthones has been reported to have an antibacterial ability, which can react with the amino acid residues on the extracellular enzymes leading to damage of proteins of biofilm matrix polymer [38,39]. Therefore, the degradation of those PLA/PBS nonwoven fabrics was decreased with MP.  Author Contributions Saowaluk Boonyod and Weraporn Pivsa-Art: Carried out experimental, analysis of PLA/PBS nonwoven fabric, summarized the literature reviews and references according to the reviewers' comments and wrote the revision of manuscript text. Phornwalan Nanthananon and Yong Ku Kwon: Carried out experimental, analysis on biodegradation of PLA/PBS nonwoven fabric, summarized the references according to the reviewers' comments, and wrote the revision of manuscript discussion.Sommai Pivsa-Art: Summarized the references according to the reviewers' comments and wrote the revision of the main manuscript text.All authors reviewed the revised manuscript (R1).

Competing Interests
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