Accepted Manuscript

Title: Chitin and waste shrimp shells liquefaction and liquefied

products/polyvinyl alcohol blend membranes

Authors: Feng-Yi Zheng, Ruisong Li, Jiadan Hu, Jie Zhang,

Xudong Han, Xinrui Wang, Wen-Rong Xu, Yucang Zhang

PII: S0144-8617(18)31278-5
DOI: https://doi.org/10.1016/j.carbpol.2018.10.079
Reference: CARP 14211

To appear in:

Received date: 20-6-2018

Revised date: 15-9-2018
Accepted date: 24-10-2018

Please cite this article as: Zheng F-Yi, Li R, Hu J, Zhang J, Han X, Wang
X, Xu W-Rong, Zhang Y, Chitin and waste shrimp shells liquefaction and
liquefied products/polyvinyl alcohol blend membranes, Carbohydrate Polymers (2018),
https://doi.org/10.1016/j.carbpol.2018.10.079

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Chitin and waste shrimp shells liquefaction and liquefied products/polyvinyl alcohol blend
membranes

Feng-Yi Zheng, Ruisong Li, Jiadan Hu, Jie Zhang, Xudong Han, Xinrui Wang, Wen-Rong Xu* ,
Yucang Zhang*

Key Laboratory of Advanced Materials of Tropical Island Resources of Ministry of Education,

College of Materials and Chemical Engineering, Hainan University, Haikou 570228, China.

Highlights:

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 Up to 92% of ball-milled chitin was liquefied by liquefaction technique.

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 Waste shrimp shells were directly liquefied and first used to modify the polymer materials.
 LBMC and LBMS obviously enhanced the mechanical property, thermal stability and

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antibacterial activity of blend membranes.
 The properties of LBMS/PVA blend membranes were comparable to those of LBMC/PVA

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blend membranes.

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Abstract: Ball-milled chitin was liquefied with an optimal yield of 92% under sulfuric acid in
diethylene glycol (DEG) at 160 °C for 120 min. The resulting liquid mixture was roughly
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separated into two portions: the real products of the reaction (liquefied ball-milled chitin, LBMC)
and the remaining unreacted DEG. LBMC was further mingled with polyvinyl alcohol (PVA) to
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prepare LBMC/PVA blend membranes. To promote the direct utilization of shellfishery waste, raw
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shrimp shells were used to replace chitin for the liquefaction and membrane preparation
operations. Liquefied ball-milled shrimp shells (LBMS) and the corresponding LBMS/PVA blend
membranes were obtained. After adding LBMC or LBMS, the mechanical, thermal, water content
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and antibacterial performance of blend membranes were significantly improved compared to pure
PVA membrane. Surprisingly, all the measured properties of LBMC/PVA and LBMS/PVA blend
membranes were comparable, and even some properties of the latter were slightly superior than
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those of the former.

Keywords: chitin; shrimp shells; liquefaction; blend membrane; biomass.

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1. Introduction
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As a widely existing renewable resource, biomass is considered as a good alternative of

petroleum for the production of fuels, high value-added chemicals and polymer materials
(Alvarez-Vasco et al., 2016). Lignocellulose consisting of cellulose, hemicellulose and lignin is
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the most abundant and low-cost biomass on the earth, and its resource utilization has attracted
great attention of scholars from different backgrounds in the past decades (Gollakota, Kishore &
Gu, 2018; Morgan et al., 2017). Among various lignocellulosic biomass conversion strategies,
liquefaction is one of the simplest and most effective techniques for converting solid biomass into
liquid polyols or aromatics (Zhao, Yan & Feng, 2013). It is usually carried out at 150–170 °C
heated temperature under atmospheric pressure using polyols as the solvent, such as polyethylene
glycol (PEG), diethylene glycol (DEG), ethylene glycol (EG) and glycol, and acids as the catalyst,

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with liquefaction efficiency from 60% to 100% (Pierson, Chen, Bobbink, Zhang & Yan, 2014).
The applications of liquefied lignocellulosic products are versatile, especially in the preparation of
polymer materials such as polyurethane foam, adhesive and epoxy resin (Pan, 2011).
In recent years, liquefaction technique is no longer limited in the plant-based lignocellulosic
materials, but has expanded to the animal cellulose, that is chitin biomass. Chitin is the second
most abundant biomass behind lignocellulose and is widely found in the exoskeletons of insects
and crustaceans. It is also the most rich nitrogen-containing polymer that consisting of N-
acetylglucosamine monomers connected by β-(1→4)-linkage in nature (Kadokawa, Shimohigoshi,
Yamashita & Yamamoto, 2015) The commercial chitin is mainly extracted from the shell waste of
the fishing industry. As reported (Yan & Chen, 2015), the annual generation of shellfishery waste,

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containing 15–40% chitin, is around 6 to 8 million metric tons globally. However, most of them

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are only regarded as rubbish to be dumped in landfills or oceans. In fact, chitin has prospective
applications in many fields, such as pharmaceuticals, textiles, cosmetics, agriculture and water

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treatment. Whereas, its application values are still underestimated due to its poor solubility in most
solvents (You et al., 2018) and the costs of extraction from shell waste (Younes, Hajji, Frachet,

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Rinaudo, Jellouli & Nasri, 2014). Making full use of chitin, or shellfishery waste without chitin
extraction, is beneficial to turn rubbish into treasure.
Yan’s group first applied liquefaction method to convert chitin into small molecular chemicals,

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including HADP, HAADP, NAG2F, DH1 etc. (Pierson, Chen, Bobbink, Zhang & Yan, 2014;
Zhang & Yan, 2016). Indeed, our group also first utilized the liquefied chitin products in the
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modification of polymer materials (Zhang et al., 2018). Ball-milled chitin was converted into
polyols in PEG 400/glycerin mixed solvents with sulfuric acid as catalyst, and the liquefaction
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yield was high to 81% at 160 ºC for 90 min. The liquefied products were further mixed with a
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polyvinyl alcohol (PVA) solution to prepare blend membranes. However, the unreacted solvents in
the liquefied products were difficult to remove due to their high boing point, and thus might affect
the characterizations of liquefied products and the performance of corresponding blend
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membranes. As is well-known, liquefaction technique can be directly applied to the conversion of

raw materials of lignocellulosic biomass, such as rice straw (Chen et al., 2018) and corn stalks
(Xu, Zhang, Ouyang, Liu & Wang, 2016). In a similar manner, can shellfishery waste be directly
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liquefied without chitin extraction and purification? If so, the applications of shellfishery waste
can become more efficient and convenient.
In the present study, ball-milled chitin (BMC) was liquefied in DEG with sulfuric acid as
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catalyst under atmosphere pressure. The optimal reaction conditions were further applied in the
liquefaction of ball-milled shrimp shells (BMS). The liquid mixture obtained by liquefaction was
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roughly separated into two portions by a rotary evaporator: the real products of the reaction,
namely liquefied ball-milled chitin (LBMC) or liquefied ball-milled shrimp shells (LBMS), and
the remaining unreacted DEG. Combined techniques incorporating Fourier transform infrared
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spectroscopy (FTIR), nuclear magnetic resonance (NMR) and gel permeation chromatography
(GPC) were applied to characterize the LBMC and LBMS. In addition, a series of LBMC/PVA
and LBMS/PVA blend membranes were prepared by a solution casting method, the latter
representing the first example of liquefied fishery shell based polymer materials. The structure and
morphology of blend membranes were investigated by FTIR, X-ray photoelectron spectroscopy
(XPS), X-ray diffraction (XRD) and scanning electron microscope (SEM) analyses. Moreover, the
mechanical properties, thermal stabilities, water content, degradation and antibacterial

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performance of blend membranes were systematically investigated.

2. Material and methods

2.1 Materials
Chitin (white powder) was purchased from Wako Pure Chemical Industries. The shrimp shells
collected from a marine food processing factory were washed thoroughly with tap water and then
dried in a vacuum oven at 60 ºC for 24 h. The dried materials were then crushed by a pulverizer to
afford the shrimp shell powder (Chen, Yang, Zhong & Yan, 2017). The chitin content in shrimp
shells powder was determined to be 29% (Ifuku & Saimoto, 2012). Sulfuric acid (98%) was
purchased from West Long Science Co., Ltd. DEG (>98%) was purchased from Shanghai Macklin

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Biochemical Co., Ltd. Ethanol (99.7%) was purchased from Guangzhou chemical reagent Co.,

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Ltd. PVA (1750±50, 99% hydrolyzed) was obtained from Sinopharm Chemical Reagent Co., Ltd.
All commercially available chemicals and reagents were used as received.

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2.2 Ball-milled chitin and ball-milled shrimp shells
Ball-milled chitin (BMC) was prepared as follows: chitin (40 g) was fully ground with a

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planetary ball mill (Fritsch, Germany) at 500 rpm using zirconia balls (diameter of 3 mm, 500 g)
in a zirconia pot (500 mL volume). The grinding process included 20 min grinding time and 40
min resting time and this operation was repeated for 6 times. After milling, the number-average

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molecular weight (Wn) and weight-average molecular weight (Mw) of chitin changed from 475520
(Da) and 489786 (Da) to 91283 (Da) and 96578 (Da) from the GPC analysis results. The
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preparation of ball-milled shrimp shells (BMS) followed the same procedure as chitin.
2.3 Liquefaction of BMC and BMS
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The liquefaction process of BMC and BMS was performed as follows: the mixture of DEG
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(28.0 g), sulfuric acid (0.4–2.0 g, 10–50 wt% of BMC or BMS) and BMC (or BMS, 4.0 g) was
intensively stirred at 160 ºC for 120 min. The reaction mixture was cooled down to room
temperature, and the solid residue was removed by filtration using a Buchner funnel and washed
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with ethanol (20 mL × 3). The resulting residue was dried in an oven at 85 ºC for 12 h.
Meanwhile, the filtrate was evaporated in a vacuum of 0.6 mbar at 100 ºC for 1 h to remove the
washing solvent (EtOH) and unreacted DEG (Budija, Tavzes, Zupančič-Kralj & Petrič, 2009), and
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the remaining liquid served as the real liquefied products, i.e., LBMC or LBMS. It was worth
noting that the unreacted DEG portion actually contained some low-boiling liquefied products and
the real products portion might contain the condensation oligomers and polymers of DEG. The
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liquefaction yield of BMC or BMS was calculated according to the following eq. (1).
𝑆𝑡𝑎𝑟𝑡𝑖𝑛𝑔 𝐵𝑀𝐶/𝐵𝑀𝑆 (𝑔)−𝑅𝑒𝑠𝑖𝑑𝑢𝑎𝑙 𝑠𝑜𝑙𝑖𝑑 (𝑔)
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Liquefaction yield (%) = × 100% (1)

𝑆𝑡𝑎𝑟𝑡𝑖𝑛𝑔 𝐵𝑀𝐶/𝐵𝑀𝑆 (𝑔)

2.4 Preparation of LBMC/PVA and LBMS/PVA blend membranes

Solution casting method was used to prepare the blend membranes. PVA (30.0 g) was
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dissolved in 600 mL of deionized water with stirring at 90 ºC for 2 h to offer a homogeneous PVA
solution. A varying percentage (0, 4, 8, 12, 16, 20 wt% to PVA) of LBMC or LBMS was added to
the PVA solution and mixed by vigorously stirring at 60 ºC for 2 h, respectively. The resulting
uniformed solution was poured into a clean glass plate and dried at room temperature for 48 h to
afford a series of blend membranes. The dried blend membranes were carefully peeled off from
the glass plate and kept in a sealed bag for further analytical use. The thickness of membrane was
measured to be about 0.09 mm.

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2.5 Structure and morphology characterization
2.5.1 Viscosity measurement
The viscosities of LBMC and LBMS were measured using a micro VISC (PheoSense, USA)
under 298.95 K and atmospheric pressure.
2.5.2 FTIR spectroscopy
FTIR spectroscopy was conducted on a TENSOR27 spectrophotometer. The test specimens
were prepared according to the regular KBr-disk method and recorded in the frequency range of
4000–400 cm–1 by co-adding 32 scans at a resolution of 4 cm–1. The deacetylation degree (DD) of
LBMC and LBMS was calculated by FTIR spectra (Abidin Munira, Junqueira‐Gonçalves Maria,
Khutoryanskiy Vitaliy & Niranjan, 2017; Baxter, Dillon, Anthony Taylor & Roberts, 1992)

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following eq. (2), where Aі is the area of the peak at wavenumber і.

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DD (%) = (100 – A1655/A3450 × 115) % (2)
2.5.3 NMR analysis
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H NMR (400 MHz) and 13C NMR (100 MHz) spectra were used to analyze the structures of

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LBMC and LBMS and were recorded on a 400 MHz spectrometer (Bruker). Chemical shift values

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were given in ppm. Residual solvent signals in the 1H spectra were used as the internal reference
(D2O: δH = 4.79).
2.5.4 GPC analysis

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GPC was applied to measure the number-average molecular weight (Mn) and weight-average
molecular weight (Mw) of liquefied products. It was performed on Styragel HT-2THF and Styragel
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HT-4THF columns using a Waters 1515 chromatographic apparatus equipped with a refractive
index detector (Waters 2414) at 45 ºC to determine the average molecular weight of LBMC and
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LBMS. A N, N-dimethylacetamide (DMAc) solution containing 0.5 wt% of LiCl was used as
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eluent and the flow rate was 1.0 mL/min (Funahashi, Ono, Qi, Saito & Isogai, 2017).
2.5.5 XPS analysis
The compositions of liquefied products and the surface of blend membranes were investigated
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by XPS (ESCALAB 250 Xi, USA) with a monochromatic Al Kα X-ray source (1486.6 eV) and
take-off angle was set at 45°. In order to compensate the surface charging effects, all bonding
energies could be used as a reference for the peak of C1s hydrocarbons at 284.6 eV. The line width
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(full width at half-maximum of Gaussian peaks or FWHM) was maintained for all components in
a particular spectrum.
2.5.6 XRD assay
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The XRD spectra of blend membranes were recorded with an X-ray diffractometer
(Panalytical XPert MPD) equipped with Cu Kα radiation and operated at 45 kV and 200 mA. The
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diffraction spectra were collected in 2θ within the range of 8–80° at a scan rate of 10° min–1.
2.5.7 SEM analysis
The blend membranes fractured by elongation at break test were used for the morphology
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investigation. The surface and cross-section of membranes were sputtered with gold, which were
observed under SEM with a Hitachi S3500N microscope operating at 10 kV.
2.6 Mechanical and thermal capacity
2.6.1 Tensile strength and elongation at break
The membranes were cut into dumbbell shapes with length 75 mm and width 4 mm. The
tensile strength and elongation at break of the blend membranes were measured using a universal
testing machine (WDW-1, Jinan, China) at a speed of 50 mm/min. Each sample was tested for 6

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times and the average and standard deviation were calculated.
2.6.2 Thermogravimetric analysis
Thermogravimetric analysis of membranes were performed using an instrument (NETZSCH-
5,209-F3) in a temperature range from 40 to 800 ºC at a heating rate of 20 ºC/min under nitrogen
atmosphere.
2.7 Water content determination
The prepared membranes with 20 × 25 mm2 excised pieces were dried in a vacuum oven at 40
ºC for 6 h and weighed (Wd). The dried samples were soaked in deionized water at 25 ºC for 72 h
(Shafiq et al., 2017). Constant weight experiments were performed to ensure that the swollen
membranes reached equilibrium. To remove excess surface water, swollen membranes were

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carefully blotted with filter paper and then weighed (Ws). The water content of the membranes was

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calculated using the following eq. (3), and the average value of three measurements was accounted
for each sample.

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𝑊𝑠 (𝑔)−𝑊𝑑 (𝑔)
Water content = × 100% (3)
𝑊𝑑 (𝑔)

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2.8 Degradation studies
To obtain the degradation profile of the prepared membranes, the samples were immersed in
buffer solutions of different pH (3.0, 7.4 and 9.0) for seven days, and the membranes were dried

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and weighted at a specific time. The loss in weight was calculated using eq. (4). Where, Wi and
Wd’ corresponded to the initial and dried weights of the membranes after immersing in different
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buffer solution, respectively.
𝑊𝑖 (𝑔)−𝑊𝑑′ (𝑔)
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Weight loss (%) = × 100% (4)
𝑊𝑑′ (𝑔)
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2.9 Antibacterial activity test

Antibacterial tests of membranes, LBMC and LBMS were separately performed against
methicillin-resistant Staphylococcus aureus ATCC 33591 (MRSA), Bacillus subtilis ATCC 6633
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(B. subtilis), Salmonella typhimurium CMCC 50115 (S. typhimurium) and Listeria monocytogenes
ATCC 19115 (L. monocytogenes)by the disk inhibition zone assay according to Pelissari’s method
(Pelissari, Grossmann, Yamashita & Pineda, 2009). The membranes were cut into 5.5 mm discs
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and placed on plates containing solid nutrient agar, and each plate was pre-coated with 0.1 mL of
bacterial inoculums. The diameters of the inhibition zones were measured with a Vernier caliper
after 24 hours of incubation at 37 ºC. Five parallel experiments were performed for each sample.
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All chemicals, glassware and other tools used were pre-sterilized at 121 ºC inside the autoclave at
a pressure of 15 psi for 30 min and then dried in an oven for later use. The experiment was
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performed on a microbial super clean bench to ensure that the whole process didn’t interfere with
the hybrid bacteria.
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3. Results and discussion

3.1 Characterization of LBMC and LBMS
The liquefaction yield of BMC raised as the amount of catalyst increased (Fig. S1). Up to 92%
of BMC was liquefied with 50 wt% of sulfuric acid in DEG at 160 °C for 120 min, while the
liquefaction yield of BMS reached 20% under the same conditions. Assuming that the LBMS was
all produced by the liquefaction of chitin in shrimp shell, the liquefaction yield could be estimated
to be 69%. Both LBMC and LBMS were shown as viscous and water soluble dark brown

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homogeneous solutions (Fig. S2), and the viscosity of LBMC (1.85 Pa·s) was obviously higher
than that of LBMS (0.47 Pa·s). FTIR investigations were conducted to compare the chemical
composition changes of BMC and BMS after liquefaction. Fig. S3 showed the FTIR spectra of
BMC, BMS, LBMC and LBMS. Overall, BMS contained BMC in its chemical composition, and
the backbones of the samples remained unchanged after liquefaction process. The detailed
assignments of BMC infrared peaks had been previously demonstrated (Pierson, Chen, Bobbink,
Zhang & Yan, 2014). The bands at about 3492 and 3262 cm–1 represented OH and NH stretching,
and the bands around 2890 and 2965 cm–1 could be assigned as CH, CH3 symmetric stretching,
and CH2 asymmetric stretching, respectively. The strong absorption peaks at 1661 to 1630 cm–1
were ascribed to amide I band, while the peaks at 1559 and 1312 cm–1 represented amide II and III

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bands. The bands of amide in the spectra of LBMC and LBMS were significantly reduced,

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indicating that some C–N bonds were broken during liquefaction. Moreover, the small absorption
occurring at 1960 cm–1 could be attributed to the –NH3+ group in LBMC and LBMS due to the

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protonation of primary amine in an acidic environment. The DD value of LBMC and LBMS was
calculated as 86.0% and 57.9%, respectively, for the baseline reference.

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The 1H NMR of LBMC and LBMS in Fig. S4a showed signals at δ = 2.23 and 2.03,
respectively, which could be attributed to the N-acetyl protons. Their 13C NMR spectra (Fig. S4b)
gave signals near δ = 27.06 and 21.90 that could be assigned as CH3 carbons of the N-acetyl

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residues, and signals at δ = 164.76 and 174.23 represented the carbonyl carbons. The signals
mentioned in LBMC were relatively weaker than LBMS, indicating that the deacetylation degree
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of LBMC was higher than that of LBMS, which was consistent with the FTIR results. Besides,
complex peaks appeared at 3.50–4.00 ppm in 1H NMR spectra and at 60.00–72.00 ppm in 13C
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NMR spectra, which could be ascribed to the methylene group from DEG derivatives, indicating
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that some of the solvent might be grafted onto the liquefied products (Chen, Chew, Kerton & Yan,
2014; Yamada & Ono, 2001).
GPC was used to evaluate the molecular weight of LBMC and LBMS. As shown in table 1, Mn
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and Mw of LBMC were 6095 (Da) and 7192 (Da), while those of LBMS were 7413 (Da) and 9711
(Da), respectively. LBMS displayed higher Mn, Mw and PDI values than LBMC, which might be
caused by the complex composition of BMS, resulting in reduced liquefaction efficiency and more
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complex liquefied products. Origin software was used for the separate the multiple peaks in the
GPC curves (Kuo, Sain & Yan, 2014). As displayed in Fig. 1, the GPC curves of LBMC and
LBMS could be divided into four peaks and five peaks, respectively, and the last peak represented
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the solvent peak. Based on the FTIR, NMR and GPC results and the previously reported
mechanism of chitin liquefaction (Pierson, Chen, Bobbink, Zhang & Yan, 2014; Zhang et al.,
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2018), a reasonable pathway was proposed for BMC and BMS liquefaction in DEG under acid
catalyst (Fig. S5).
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3.2 Structure and morphology of blend membranes

3.2.1 FTIR analysis
FTIR spectra of LBMC/PVA and LBMS/PVA blend membranes with different content of
liquefied products (0, 4, 8, 12, 16, 20 wt%) were shown in Fig. 2. The pure PVA membrane
showed the characteristic absorptions at 3443 cm–1(OH), 2925 cm–1(CH2) and 1095 cm–1 (CO)
(Deng, Li, Yang & Li, 2014) in Fig. 2a and 2b. The blend membranes presented the absorptions of

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both PVA and liquefied products. Compared with PVA membrane, it was clearly noticed that a
new peak (1126 to 1060 cm-1) that represented the C-O stretching vibration and asymmetric bridge
oxygen (Pearson, Marchessault & Liang, 1960) in the blend membrane, which came from the
LBMC or LBMS. The width change of ~3443 cm–1 suggested a possible overlapped stretching
between the –NH2 (in LBMC or LBMS) and the –OH (in PVA) groups. In addition, the
disappearance of absorptions at 1960 cm–1 which attributed to the –NH3+ group suggesting some
physical and chemical interactions between PVA and LBMC (Sharma et al., 2016).

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3.2.2 XPS results

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XPS analysis was performed to quantitatively evaluate the chemical composition of liquefied
products and the membrane surface. The main elements contained in the measured samples were

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carbon (C), nitrogen (N) and oxygen (O), as shown in the XPS spectra in Fig. 3 (left). The N
content in LBMS and LBMS/PVA blend membrane was significantly higher than that in LBMC

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and LBMC/PVA blend membrane, which might be caused by the liquefaction of some proteins in
the raw shrimp shells. Table S1 listed the chemical composition of liquefied products and the
surface of blend membranes obtained from the XPS analysis. Furthermore, the C1s peak was

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scanned in detail to detect the deconvolution in Fig. 3 (right). The binding-energy peaks had been
identified as CH2 or C-H bonds at 284.6 eV, C-N at 286.0 eV and N-C=O at 288.4 eV (Huang,
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Fang, Chen, Lu & Zhang, 2017; Miao, Zhang & Bai, 2015). The distribution of N-C=O in LBMC
was obviously lower than that of LMBS, indicating that the deacetylation degree of LBMC was
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higher than that of LBMS. Not surprisingly, this result was again in accord with the FTIR and
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NMR results.
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3.2.3 XRD Characterization

The XRD patterns of LBMC/PVA and LBMS/PVA blend membranes were presented in Fig. 4.
It can be seen that the peak occurring at about 2θ = 19.8° corresponded to the 101 plane in the
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PVA membrane, which was chiefly due to the intramolecular hydrogen bonds formed between the
–OH groups on the PVA skeleton (Zhao, Qian, An, Zhu, Yin & Sun, 2009). Apparently, with the
increase of LBMC or LBMS content, a new peak appeared at around 2θ = 12.3° and gradually
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enhanced, which might be the characteristic peak of LBMC and LBMS (Zhang, Zou, Wei, Mu,
Liu & Tong, 2017). With the addition of liquefied products, the peak intensity at 2θ = 19.8°
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increased first and then decreased, and the peak intensity reached a maximum when the
concentration of liquefied product was 12%. These results indicated that the crystallinity of PVA
membrane was influenced by LBMC or LBMS content in blend membranes. The plausible reason
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was that moderate liquefied products acted as nucleation sites for the crystallization of PVA
(Cadek, Coleman & Barron, 2002), resulting in the increase of crystallinity. The further decrease
in crystallinity of blend membranes was probably because the excess liquefied products caused the
decrease of intramolecular hydrogen bonding in PVA moiety and increase of intermolecular
interactions between PVA and liquified products (Lu, Peng, Jiang & Wang, 2006; Chen & Chen,
1996). The crystallinity decrease tendency of LBMS/PVA blend membranes appeared more
obvious, which presumably due to the more complicated components of LBMS. .

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3.2.4 Scanning electron microscopy
The surface and cross-section SEM images of pure PVA membrane and selected blend
membranes were presented in Fig. 5. Particles appeared on the surfaces of LBMC/PVA blend
membranes, indicating partial phase separation behavior. As the concentration of LBMC increased
to 16%, the phase separation behavior was more pronounced. While, the LBMS/PVA blend
membranes had smooth surfaces and compact textures. No signs of phase separation were
observed, nor irregularities such as air bubbles, pores or cracks, indicating that LBMS and PVA

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were highly compatible (Liu, Wang & Lan, 2018). The results suggested that LBMS was more

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compatible with PVA than LBMC.

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3.3 Properties of blend membranes

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3.3.1 Mechanical property
The tensile strength and elongation at break of pure PVA, LBMC/PVA and LBMS/PVA blend
membranes were measured. As shown in Fig. 6, incorporating LBMC and LBMS significantly

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improved the mechanical properties of the PVA membrane, which might be attributed to the
formation of strong intermolecular interactions between liquefied products and PVA polymers
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(Deng, Li, Yang & Li, 2014). The partial deacetylated chitin in LBMC and LBMS gave –NH2
groups that further interacted with –OH groups in chitin and PVA, leading to the formation of
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strong polymer networks (Sharma et al., 2016). Surprisingly, the effect of LBMS addition on the
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performance of PVA membrane was even better than that of LBMC. In general, mechanical
properties enhance as the molecular weight increases (Nunes, 1982). Therefore, this was probably
due to the higher molecular weight of LBMS as evidenced by GPC characterization. The behavior
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accounted for the tensile strength initially increased and then decreased, and the elongation at
break gradually enhanced with the increase of the addition amount of LBMC or LBMS. In
general, when the concentration of LBMC or LBMS was 12 wt%, the tensile strength and
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elongation at break reached the optimal values, i.e. 59.5 MPa and 701%, 67.15 MPa and 759%,
respectively for LBMC/PVA and LBMS/PVA blend membranes. Stress-strain curves of blend
membranes were displayed in Fig. S6. The mechanical performance results were in line with the
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XRD results, indicating that the crystalline structure of the membrane was beneficial to enhance
its mechanical properties.
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3.3.2 Thermogravimetric analysis

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TGA and DTG analyses were carried out to detect the thermal stability of the blend
membranes. The degradation occurred in three well-differentiated steps for LBMC/PVA blend
membranes (Fig. 7a and c): (i) the volatilization of water and oligomers at 40–160 ºC; (ii) the
decomposition of LBMC and PVA side chains at 160–350 ºC; (iii) the breakage of the LBMC and
PVA backbones at 410–500 ºC. In contrast, the main weight loss was the latter two stages. The
TGA data in table S2 showed that both the maximum degradation temperature (Tmax) and the
statistic heat-resistant index temperature (Ts) of the LBMC/PVA blend membranes increased first

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and then decreased with increasing LBMC content. When the concentration of LBMC was 4%,
the Ts (110.3 ºC) and Tmax (308 ºC, 455 ºC) of the blend membrane reached the maximum value,
which was much higher than the Ts (93.7 ºC) and Tmax (266 ºC，443 ºC) values of PVA
membrane. Unlike the LBMC/PVA blend membrane, only two weight losses were observed in the
TAG and DTG curves of LBMS/PVA blend membranes (Fig. 7b and d). The weight loss at 40–300
ºC was probably attributed to the moisture vaporization and the side chain decomposition of
LBMC and PVA (Lee, Lopez Hernandez & Moore, 2015). The other weight loss at 300–495 ºC
corresponded to the degradation of LBMS and PVA backbones (García-Cruz, Casado-Coterillo,
Iniesta, Montiel & Irabien, 2016). It was obvious that the thermal stability of the membrane was
greatly improved after the addition of LBMS to the PVA membrane. In particular, the Tmax of 12%

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LBMS/PVA membranes reached 293 ºC and 448 ºC, which was superior than that of PVA

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membrane (266 ºC and 443 ºC, Table S3). These results showed that the addition of LBMC or
LBMS was beneficial to enhance the heat resistance of the blend membranes, and the effect of

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adding LBMS was more significant.

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3.3.3 Water content

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The water content of the pure PVA and blend membranes was displayed in Fig. S7. As the
amount of liquefied products increased, the water content of the blend membranes gradually
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increased. This was due to the hydrophilic nature of LBMC and LBMS which could offer
considerable hydrogen bonding with the hydroxyl groups of PVA (Shafiq et al., 2017). The
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improved swelling performance of blend membranes indicated that they had more flexible
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structures and higher fluxes.

3.3.4 Degradation studies
The membranes were immersed into a buffer solution of different pH (3.0, 7.4 and 9.0) to test
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the degradation behaviors. The weight loss data in Fig. 8 revealed that the addition of liquefied
products gently impeded the degradation of blend membranes under different pH conditions, but
still remained an acceptable level. This was mainly because that the hydrogen bonded networks
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between liquefied products and PVA prevented the hydrolytic degradation of blend membranes
(Liu, Wang, Zhang, Liu, Wang & Song, 2015). In addition, low pH condition was more conducive
to the degradation of blend membranes because the acid could weaken the interaction between
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liquefied products and PVA, and promote hydrolysis of polymer chains.

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3.3.5 Antibacterial activities

The antibacterial activities of LBMC, LBMS and 12% blend membranes were tested against
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four microorganisms (MRSA, B. subtilis, S. typhimurium and L. monocytogenes ) by disk method.

As shown in Fig. 9, LBMC and LBMS appeared excellent antibacterial activities in all four
microorganisms. The diameters of the inhibition zones were listed in Table S4. As expected,
LBMC showed slightly better inhibition than LBMS because only 129% of chitin was measured in
shrimp shells. According to previous studies (Yang, Chou & Li, 2005), the antibacterial activity of
liquefied products was supposed to be attributed to their abundant protonated amine groups in the
structure. It can be seen that the blending of LBMC or LBMS with PVA greatly improved the

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antibacterial activities of the membranes. Moreover, the inhibition zones of L. monocytogenes
(Gram-positive bacteria) were slightly larger than S. typhimurium (Gram-negative bacteria). This
phenomenon might be related to the extra outer membrane surrounding the cell wall of Gram-
negative bacteria, which limited the diffusion of the hydrophobic compounds through the
lipopolysaccharide coating (Burt, 2004). The suspected bacteriostatic mechanism was that the
active ingredients in liquefied products and the corresponding blend membranes attacked the
phospholipid membrane of the cell, resulting in the increased cytoplasm permeability and leakage,
or the interaction with enzymes located on the cell wall (Perumal, Sellamuthu, Nambiar & Sadiku,
2018; Zivanovic, Chi & Draughon Ann, 2006).

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4. Conclusions
Ball-milled chitin and ball-milled shrimp shells were effectively converted into liquids in DEG

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by liquefaction technique. FTIR, NMR and GPC analyses of LBMC and LBMS indicated that
deacetylation and depolymerization reactions occurred during liquefaction process, and the DD,

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Mn and Mw of LBMC and LBMS were calculated as 86.0% and 57.9%, 6095 (Da) and 7413 (Da),
7192 (Da) and 9711 (Da), respectively. In addition, the blending of LBMC or LBMS significantly
enhanced the mechanical and thermal properties of PVA membrane, which might be contributed to

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some chemical and physical interactions between –OH, –NH2 and –NHAc groups in LBMC or
LBMS and –OH groups in PVA. Besides, the water content and antibacterial activity of blend
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membranes were notably improved. Although the degradation rate was slightly decreased, it still
remained at an acceptable level. Most of all, the properties of LBMS/PVA blend membranes were
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comparable to those of LBMC/PVA blend membranes, which would promote the direct utilization
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of shellfishery waste.
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Acknowledgements
We gratefully acknowledge the support from the Key Scientific Research Project Funding of
Hainan Province (grant number ZDYF2017005) and Hainan Natural Science Foundation (grant
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number 217008).
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Fig. 1. GPC analysis of LBMC (a) and LBMS (b).

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Fig. 2. FTIR spectra of 0–20 wt% LBMC/PVA blend membranes (a)

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and 0–20 wt% LBMS/PVA blend membranes (b).

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Fig. 3. X-ray photoelectronic spectrum (left) and deconvolutions of C1s (right) of LBMC (a),

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LBMS(b), 12% LBMC/PVA (c), 12% LBMS/PVA (d) blend membranes.

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Fig. 4. XRD spectrums of LBMC/PVA (a) and LBMS/PVA (b) blend membranes with different

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liquefied products content (0, 4, 8, 12, 16, 20 wt%).

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Fig. 5. SEM images of the surfaces (a–g) and cross-sections (h–n) of PVA (a, h),
8% LBMC/PVA (b, i), 12% LBMC/PVA (c, j), 16% LBMC/PVA (d, k),

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8% LBMS/PVA (e, l), 12% LBMS/PVA (f, m) and 16% LBMS/PVA blend membranes (g, n).

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Fig. 6. The effect of the LBMC and LBMS on the mechanical capability of the blend membranes.

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Fig. 7. TGA (a) and DTG (c) curves of LBMC/PVA blend membranes, TGA (b) and DTG (d)
curves of LBMS/PVA blend membranes.

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Fig. 8. Degradation studies of LBMC/PVA and LBMS/PVA bend membranes at pH 3.0, 7.4 and
9.0.

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Fig. 9. Antibacterial activity of PVA, 12 wt% LBMC/PVA, 12 wt% LBMS/PVA, LBMC and

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LBMS against bacterium.

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Table 1
GPC data of LBMC and LBMS.
Codes Mn Mw PDI Peak 1 Peak 2 Peak 3 Peak 4
LBMC 6095 7192 1.18 17906 6310 4112 —
LBMS 7413 9711 1.31 14927 9661 8208 3820

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