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PII: S0141-8130(20)33277-3
DOI: https://doi.org/10.1016/j.ijbiomac.2020.05.122
Reference: BIOMAC 15605
Please cite this article as: W. Youzhi, W. Jincheng, S. Shiqiang, et al., Preparation and
application properties of sustainable gelatin/chitosan soil conditioner microspheres,
International Journal of Biological Macromolecules (2020), https://doi.org/10.1016/
j.ijbiomac.2020.05.122
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* Corresponding author: wjc406@sues.edu.cn; Tel.: 8621-67791219; Fax: 8621-67791214
ABSTRACT: Traditional soil conditioners can be used as drugs to regulate elements in soil due to
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overuse or pollution. There are some negative effects such as low efficiency, hard to degrade and harmful
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to the environment when they are used. Soil conditioners prepared from sustainable macromolecules can
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reduce or eliminate these negative effects. In this work, gelatin (GL), chitosan (CS), magnesium
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ammonium phosphate (MAP) and fulvic acid (FA) were combined and prepared into GL/CS/MAP-FA
soil conditioner microspheres with core-shell structure through emulsion crosslinking method. Scanning
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electron microscopy (SEM) and Fourier infrared spectroscopy (FT-IR) analysis results clarified the
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surface composition of the microspheres and revealed the good coating performance of GL/CS shell
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materials. The thermal stability of the microspheres was measured by thermogravimetric analysis (TGA).
Water-retention, degradation and crop cultivation showed that these microspheres owned good
biocompatibility, which can extend the water-retention time of the soil, settle free heavy metal ions and
promote crop growth. The load of fertilizer in the microsphere can effectively prolong the release cycle
1. Introduction
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Although significant progress has been made in crop varieties, fertilizers, pesticides and land
management methods, there has been a downward trend in agricultural production, which is inextricably
linked to current soil problems [1]. Studies have shown that soil erosion and degradation are occurring at
an unprecedented rate in industrialized and developing countries, with far-reaching implications for the
nature of soil ecosystems [2]. In order to improve the yield and quality of agricultural products, we must
Scholars at home and abroad have used a variety of methods to improve soil quality. For instance, lime
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was applied to regulate soil acidity; natural zeolite was used to protect nitrogen fertilizers and to adsorb
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heavy metal ions from soil; carbon and humus were used to increase the content of organic matters in the
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soil; natural macromolecular or synthesized polymer hydrogels were used as water holding agents [4].
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However, these methods may bring some problems to the soil. Lime may cause aggregation for soil
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particles, and this may lead to soil compaction [5]; natural zeolite owned low utilization efficiency, and it
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was costly to improve utilization efficiency through artificial modification [6]; aged biochar has a
negative impact on earthworms and soil microorganisms, inhibits the growth of some plant roots, reacts
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with nutrients in the soil, and also affects the decomposition of some organic matter [7]; polymers such as
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polyacrylamide are difficult to degrade, and small molecules generated during degradation are harmful to
Above soil conditioning practice shows that the existing soil conditioners may have some advantages
but are difficult to meet the requirements of people, such as environmental friendliness, intelligent
response, controlled release, sustainability and high efficiency. That is, they cannot meet the future
development trend of soil conditioners. Since the beginning of the 21st century, the sustainability of
human production activities and the reduction of carbon dioxide emissions have attracted much attention.
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Thus, research on environmental protection, resources and energy saving and resource regeneration has
been intensified. Therefore, researchers in the world have made great efforts to develop and utilize
Naturally derived macromolecules usually have good water absorption and sustainability, and also have
many advantages such as high natural abundance, renewability, and low cost [9]. It’s a good choice to
apply it to soil conditioner or fertilizer carrier. Gelatin (GL) and chitosan (CS) have good biocompatibility
and sustainability, and can be miscible at any ratio through intermolecular hydrogen bonding and
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electrostatic interaction [10, 11]. As a type of traditional soil conditioner, fulvic acid (FA) can improve
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drought tolerance, regulate nutrient ingestion, stabilize pH values of soil, and decrease fertilizer extracting
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behaviors [12]. For magnesium ammonium phosphate (MAP), another type of soil conditioner, it is rich
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in N, P and Mg and has good biocompatibility [13]. It can be used as a slow-release fertilizer, and can
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also react with heavy metals such as Pb and Cu in the soil to form highly insoluble metal-bearing
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hydroxyapatites [14]. When above materials were combined into microspheres or microcapsules with a
In this study, GL and CS microspheres were prepared by the emulsion crosslinking method, and FA
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together with MAP were inside. A series of experiments such as swelling, water-retention, slow release
and degradation were used to characterize their structure and properties. Results showed that these
microspheres possessed certain water-retention capacities, sustained release ability, good degradation
2. Experimental
2.1 Materials
Liquid paraffin, acetic acid (99% pure), gelatin (GL, type B), chitosan (CS, 90%+(deacetylated),
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relative molecular weight M n 52844 g / mol , PDI = 1.057), glutaraldahyde (25% in water) and copper
sulfate pentahydrate (CuSO ·5H2O) were purchased from Shanghai Titan Scientific Company (Shanghai,
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China). Fulvic acid (FA, 90% pure) was received from Cool Chemistry (Beijing, China). Sorbitan
monooleate (Span 80) and sodium diethyldithiocarbamate trihydrate (DDTC) were obtained from
Macklin Chemical (Shanghai, China). Magnesium ammonium phosphate (MAP) was self-made by
precipitation.
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The microspheres were prepared by a modified emulsion crosslinking method [16]. 15.0 g of GL was
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dissolved in 100.0 ml of deionized water at 50 °C and 3.0 g of CS was dissolved in 100.0 ml of dilute
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acetic acid solution (volume concentration, 2%) at room temperature. 0.5 g of FA and 1.0 g of MAP were
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added to the beaker, followed by the addition of 10.0~15.0 ml of GL and 5.0~10.0 ml of CS solutions,
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respectively. A three-necked flask was poured with 250.0 ml of liquid paraffin, followed by 1.0 ml of
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emulsifier Span 80 and then crosslinked with 1.0 ml of glutaraldehyde solution (volume concentration,
0.5%). Finally, a type of sustained release microspheres, GL/CS/MAP-FA, were obtained after
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centrifuged at low speed, washed with petroleum ether, filtered, and dried at room temperatures.
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2.3 Characterization
Polarized optical microscope (POM) was used to observe the overall morphology of GL/CS/MAP-FA
microspheres. They were first dispersed with water, and a small amount of this mixture was taken and
dropped on a glass slide. Then, this mixture was observed by POM after excess water was removed with
absorbent paper. Scanning electron microscopy (SEM) was applied to observe surface morphology of the
microspheres using a Hitachi S4800 SEM. This equipment was operated at an accelerating voltage of 5
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kV. Before the observation, the microspheres adhered to the conductive adhesive were sprayed with gold
powders.
The crosslinking bonds generated by GL/CS/MAP-FA microspheres were measured by a Nicolet FT-
IR380. The raw material was pressed into a flake and then detected by Attenuated Total Reflectance
(ATR) method.
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TGA curves of the microspheres were obtained by Linsesis STA PT-1000. The microspheres were
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added to a crucible, and the heating rate was set to 10 °C/min. The samples were then heated to 600 °C
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under a N2 atmosphere.
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2.4 Swelling and water-retention behavior of GL/CS/MAP-FA microspheres
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The swelling test method was adopted from gravimetric method [16]. 0.2 g of GL/CS/MAP-FA
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microspheres was added into a 50.0 ml of centrifuge tube, and the original weight m0 was recorded. All
centrifuge tubes were numbered and 20.0 ml of deionized water or phosphate buffer solution (pH=4.6)
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was added. At intervals, one sample was taken from each group, filtered to remove water, weighed, and
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mt m 0
Swelling ratio 100%
m0
The water-retention test was adopted from a previous investigation [17]. Ten paper cups were selected
and holes were punctured at their bottoms. Then, they were numbered separately. 0.4 g of GL/CS/MAP-
FA microspheres was weighed and put into the corresponding paper cup, respectively. Then, 40.0 g of
sand were added, weighed and recorded as m0 . 50.0 ml of deionized water were poured into all paper
cups, and stood until no water seeped out at the bottom of all the paper cups. Then, the system was
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weighed and ms was recorded. Finally, the paper cups were placed in an oven at 30 °C, and they were
mt m 0
Water retention ratio 100%
m s m0
The FA sustained release behavior was conducted using the membrane dialysis method [18]. Before the
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0.3 g of CL/CS/FA-MAP microspheres was added in a dialysis bag, sealed and placed in a 100 ml of
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centrifuge tube. One group was added with 50.0 ml (V0) of deionized water (pH=7.0), and the other group
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was added with 50.0 ml of a phosphate buffer solution (pH=4.6). The sample was placed at room
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temperature and 2.0 ml of sustained-release solution was taken out and diluted to 30.0 ml at intervals.
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Then, 2.0 ml (V1) of blank solution was added to the centrifuge tube. The absorbance was measured at
271 nm and the solution concentration of each sample was calculated. The total amount of FA released at
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each time point is equal to the amount of FA in the centrifuge tube solution plus the amount of FA in the
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removed solution.
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Standard curves:
At 4.8498 C t R 0.9978
2
Total release:
M t V0 C t 0 C t 1 V1
t 1
Mt
FA release ratio 100%
Mq
At: the value of the absorbance measured at time t; Ct: the concentration of FA corresponding to the
absorbance measured at time t; V0: the volume of solution initially added to the centrifuge tube; V1: the
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volume of sustained release solution take out each time; Mt: the total mass of FA released by
M t M kt1/2 (2)
M t M kt n (3)
M t ( M t ) kt 1/2 b (4)
Mt/M∞: the release ratio of FA at time t; k, b: kinetic constants; n: diffusion exponent (n≤0.43, Fickian
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diffusion; 0.43 < n < 1.0, non-Fickian or anomalous transport).
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2.6 Heavy metal adsorption behavior and kinetics of GL/CS/MAP-FA microspheres
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Copper sulfate was used as a heavy metal source to test the fixed effect of microspheres on heavy metal
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ions. 150.0 ml (V0) of copper sulfate solution (5 mmol/L) was added into 250.0 ml of conical bottle, and
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0.5 g of CL/CS/FA-MAP microspheres was added at intervals. The sample was then processed with
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DDTC, and the sample absorbance was tested at 448 nm. Thus, the fixed amount of copper ions can be
calculated.
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Standard curves:
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At 5.4496 C t R 0.9991
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Total fixation:
M t M 0 (Ct V0 0 Ct 1 V1 )
t 1
Mt
Total fixation ratio 100%
M0
At: the value of the absorbance measured at time t; Ct: the concentration of Cu2+ corresponding to the
absorbance measured at time t; Mt: the total mass of CuSO4 fixed by microspheres at time t; M0: the total
mass of CuSO4 contained in copper sulfate solution; V0: the total volume of solution; V1: the volume of
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k1t
ln(qe qt ) ln qe (6)
2.303
t 1 t
2
(7)
qt k2 qe qe
k1, k2 and kint: kinetic constants; C: the boundary layer thickness; qt, qe: removal capacity at time t and
equilibrium, respectively.
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2.7 Degradation and biological toxicity performance of GL/CS/MAP-FA microspheres
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The samples were degraded in deionized water (pH=7.0) and phosphate buffer solution (pH=4.6),
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respectively. Each group of samples was weighed 0.1±0.01 g, and the initial value w0 was recorded. Then,
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it was placed into a 5.0 ml of centrifuge tube containing 4.0 ml of deionized water or phosphate buffer
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solution. The samples were taken from a centrifuge tube for each measurement, and then dried, weighed,
and recorded as wt. The weight loss ratio was calculated as follows:
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w0 wt
Weight loss ratio 100%
w0
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In order to test the biotoxicity of microspheres, microbial culture was carried out in accordance with
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previous study [19]. A culture medium with the preparation materials of microspheres was sterilized
under an ultraviolet lamp, and was exposed to the air for 24 hours. Then, the medium was transferred into
the incubator and the growth of the mold on the medium was observed.
For the plant test of garlic, twelve PVC pipes with length of 30 cm were divided into four groups, and
were then numbered separately. They were filled with a sufficient amount of sand, and added with 0.5 g
of GL/CS/MAP-FA microspheres. The corresponding PVC pipe was buried in the sand with depth of 5
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cm. Then, the treated garlic seedlings were planted in the pipe and their growth status was recorded.
Later, wheat, chrysanthemum coronarium and corn were planted under the same experimental conditions.
The schematic diagram for preparation and function of GL/CS/MAP-FA microspheres was presented in
Fig.1. First, the core materials, FA and MAP, were added to the GL solution. Then, the CS solution was
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poured into and was evenly mixed. The liquid paraffin was added into the mixture, and was emulsified
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into spherical droplets by mechanical shearing. Last, the crosslinking agent, glutaraldehyde, was added,
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and its -CHO groups may react with -NH2 groups in GL and CS to form C=N bonds [20]. Thus, a three-
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dimensional network structure with a microsphere shape, GL/CS/MAP-FA microsphere, was finally
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formed.
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After the microspheres were applied to the soil, FA on the surface of microspheres could be quickly
diffused to the soil. In addition, MAP could also react quickly with heavy metal cations in the soil,
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generating insoluble heavy metal phosphate and releasing N and Mg elements needed for plant growth.
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After a certain period of time, microspheres began to degrade in the soil, and FA and MAP in the
microsphere fragments were continuously released into the soil until the microspheres were completely
degraded.
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Fig.1 Schematic diagram for preparation and function of GL/CS/MAP-FA microspheres.
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Fig.2 (a), (b): POM pictures of GL/CS/FA microspheres without adding MAP before (a) and after (b) drying;
(c), (d): POM pictures of GL/CS/MAP-FA-2 microspheres before (c) and after (d) drying;
(e)-(g): SEM pictures of GL/CS/MAP-FA microspheres with a magnification of 150x(e), 500x(f), and 2000x(j);
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(h): SEM picture of MAP in GL/CS/MAP-FA microspheres; (i)-(l) SEM pictures of GL/CS/MAP-FA microspheres profile
with a magnification of 15000x(i), 25000x(j), 35000x(k) and 110000x(l).
Table 1. The amount of the drug used in the preparation of GL/CS/MAP-FA microspheres
Samples GL/ml (15 g/100ml) CS/ml (3 g/100ml) FA/g MAP/g
GL/CS-FA 10.0 10.0 0.25 0
GL/CS/MAP-FA-1 15.0 5.0 0.25 1.0
GL/CS/MAP-FA-2 12.5 7.5 0.25 1.0
GL/CS/MAP-FA-3 10.0 10.0 0.25 1.0
The amount of the drug used in the preparation of GL/CS/MAP-FA microspheres was summarized in
Table 1. The three samples were similar in macroscopic and microscopic morphology. Fig.2 presented the
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Before drying, the surface of GL/CS-FA microspheres without MAP (Fig.2(a)) exhibited no significant
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difference from that of GL/CS/MAP-FA-2 microspheres (Fig.2(c)). After drying, GL/CS/MAP-FA-2
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microspheres exhibited no obvious defects (Fig.2(d)) and GL/CS-FA showed obvious collapse (Fig.2(b)).
Here, MAP plays a supporting role and avoids the collapse of GL/CS/MAP-FA microspheres. SEM of the
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microspheres after drying was given in (e)-(l), respectively. In the microscopic state, it can be seen that
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the surface of the microspheres was uneven and many powders were embedded. In addition, some small
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pores can be seen on the surface of the microspheres. The MAP with the porous flower-like structure
(Fig.2(h)) and the network-like pore structure (Fig.2(i-l)) inside the microsphere can be seen from the
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sectional view of the microspheres. The microspheres were filled with interconnected micropores, and
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water can flow freely from these holes [21]. The presence of surface and internal holes allowed water and
air to enter the interior of microspheres, which facilitated rapid water absorption and oxidative
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The main composition of raw materials and different microspheres can be deduced from FT-IR analysis
(Fig.3(a)). The absorption peak at 3300 cm-1 was referred to the stretching vibration of –OH, -NH2 and
NH4 groups in CS, GL and MAP, and the peak at 2930 cm-1 was the absorption peak of C-H bonds. The
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characteristic absorption peaks at 1635, 1540 and 1240 cm-1 were ascribed to the stretching vibrations of
the C=O, N-H and C-N bonds, respectively. In addition, a characteristic absorption peak for the stretching
vibration of C=N bond was shown at 1750 cm-1, which was produced by a Schiff base reaction between -
CHO groups in glutaraldehyde and -NH2 groups in GL and CS [20]. The absorption peaks of GL/CS and
GL/CS/MAP-FA microspheres were basically the same, and the characteristic peaks of FA at 3400, 1400
and 1080 cm-1 were not visible in the infrared spectrum of GL/CS/MAP-FA microspheres. This may be
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The thermal degradation process of the microspheres was presented in Fig.3(b) and divided into three
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stages [22]. The first stage was approximately from 30 to 190°C, and this was ascribed to the continuous
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vaporization of the moisture absorbed and bounded in the samples. In the range of 70-190°C, MAP would
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gradually undergo a thermal decomposition reaction [23]. Therefore, a peak at 113°C was exhibited in the
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DTG curves. In the range between 190 and 400°C, especially at 314°C, weight loss may be caused by the
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dehydration of protein and cleavage of C-O-C bonds in the polymer backbones and carboxyl groups. In
addition, the slow weight loss in the range from 400 to 600°C was attributed to the gradual destruction of
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the crosslinking points and the further cleavage of the remaining organic matters.
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Fig.4 (a): Swelling behavior of GL/CS/MAP-FA microspheres under neutral and acidic conditions;
(b): Water-retention behavior of GL/CS/MAP-FA microspheres.
Fig.4 presented swelling and water-retention behaviors of GL/CS/MAP-FA microspheres under neutral
and acidic conditions. The degree of gel swelling is related to the pore of the polymer network and the
interaction between polymer and solvent [24]. As one composition of the microsphere, GL shows the best
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shown in Fig.4(a), under neutral conditions, the order for the water absorption amount of the
gel volume is also influenced by environmental factors [25]. Under acidic conditions, the swelling ability
of GL/CS/MAP-FA microspheres is also affected by factors such as solution concentration and solution
acidity. The water absorption of CS in microspheres also increased, resulted in a change in the ranking of
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estimation showed that these curves approached the logarithmic and power function curve growth mode.
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The swelling ratio of microspheres was also related to factors such as the solution environment and the
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type of microspheres, and thus a difference was presented in the degree of curve fit. From Fig.4(b), the
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water loss ratio of the microspheres was positively correlated with the water absorption capacity of the
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microspheres. However, compared to the blank group, the water retention capacity of the experimental
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group with microspheres was not ideal. Compared with the hydrogel prepared by Chen et al. [8], the
cross-linking point limited the water absorption and the dispersibility of GL/CS/MAP-FA microspheres,
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3.5 Sustained release behavior and kinetic of GL/CS/MAP-FA soil conditioner microspheres
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Fig.5 (a): Sustained release behavior of GL/CS/MAP-FA microspheres under neutral and acidic conditions.
(b): Linear relationship of GL/CS/MAP-FA microspheres under mathematical model.
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Table 2. Parameters of sustained release kinetic analysis of GL/CS/MAP-FA microspheres according to different models.
Models
Code first-order Higuchi Ritger-Peppas parabolic diffusion
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2 2 2
k R k R k n R k R2 b
GL/CS/MAP/FA-1
0.01115 0.81883 0.03138 0.82401 0.319979 0.20354 0.90069 0.19398 0.98949 -0.00943
pH=7.0
GL/CS/MAP/FA-2
0.01411 0.97545 0.03525 0.83106 0.287927 0.22935 0.92436 0.18156 0.99235 -0.0083
pH=7.0
GL/CS/MAP/FA-3
0.01581 0.95029 0.03491 0.72401 0.273345 0.24544 0.85027 0.16383 0.97783 -0.00618
pH=7.0
GL/CS/MAP/FA-1
0.01331 0.94669 0.03798 0.8168 0.220112 0.27822 0.88063 0.13804 0.96429 -0.00443
pH=4.6
GL/CS/MAP/FA-2
0.01397 0.93205 0.03769 0.82498 0.233732 0.26735 0.88251 0.14714 0.95843 -0.0052
pH=4.6
GL/CS/MAP/FA-3
0.01832 0.91508 0.0379 0.83363 0.235701 0.26629 0.88757 0.14837 0.9634 -0.00529
pH=4.6
The sustained release profiles of GL/CS/MAP-FA microspheres were presented in Fig.5(a). After
reaching equilibrium, the FA in the microspheres was basically released into the sustained release
solution. Based on the total amount of FA released from the microspheres, the drug-loading of
GL/CS/MAP-FA-1,2,3 microspheres can be calculated as about 10%, 7% and 7%, respectively. Under
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neutral condition, GL/CS/MAP-FA-2,3 microspheres with less drug loading first reached the release
equilibrium, and GL/CS/MAP-FA-3 microspheres owned the shortest release equilibrium time. However,
GL/CS/MAP-FA-1 microspheres exhibited sustained release behavior and the longest equilibrium time
due to higher drug loading. Under acidic condition, the release equilibrium time of GL/CS/MAP-FA-3
microspheres was still the shortest, and the gap with the GL/CS/MAP-FA-1,2 microspheres was
significantly shortened. The time for GL/CS/MAP-FA-1,2 microspheres to reach the sustained release
equilibrium was basically the same. Factors such as pore diffusion, degradation, and surface desorption of
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polymeric microcapsules may affect the sustained release behavior of the microcapsules [26]. Within 24
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h, the FA in the microsphere was released quickly. This was caused by the rapid dissolution of the FA on
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the surface of microspheres, which facilitated the rapid action of the microsphere on the soil after
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application. The sustainable release data were subjected to curve regression analysis, and the logarithmic
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estimated curves (R2> 0.90) had the best fit with the measured value.
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In order to study the release behavior of microspheres, four mathematical models, first-order, Higuchi,
Ritger-Peppas and parabolic diffusion (Equation (1)-(4)), were used to describe the sustained release
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process of FA [27]. The first-order model is often used to explain the release process including ion
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exchange and dissociation, and the dissolution rate is determined by drug content [28]. The Higuchi
model is applied to describe the dissolution process of Fickian diffusion as a rate-limiting step [29]. In
addition, Ritger-Peppas model is normally applied to explain drug diffusion and dissolution, and the
parabolic diffusion model is suitable for elucidating the release process of outer surface/edge diffusion as
Based on the kinetic models, some parameters were calculated and given in Table 2. It can be seen
that R2 values of the sustained-release data by the four mathematical models were quite different, and this
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value was the closest to 1 in the parabolic diffusion model (R2 > 0.95). The closer the R2 value is to 1, the
better the model fits the sustained release data. As can be seen in Fig.5(b), most of the fitted data of the
parabolic diffusion model and the first-order model were connected into a straight line, and the remaining
points were also closely scattered on both sides of the straight line. The fitted data of the Higuhi and
Ritger-Peppas models rose in a parabolic trend, and most of the data points were scattered outside the
fitted straight line. This showed that the sustained release process of microspheres was more complicated.
FA diffused into the solution from the surface of GL/CS/MAP-FA microspheres, and the controlling step
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was the outer surface/edge diffusion process, which was accompanied with ion exchange and dissociation
process.
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3.6 Heavy metal removal capacity and kinetic of GL/CS/MAP-FA soil conditioner microspheres
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Fig.6 (a): Cu2+ removal capacity for different samples; (b): Cu2+ removal capacity for different samples within 0-35 hours;
(c): Full-scan graph after coloring agent is added to different concentrations of Cu2+ solution; (d)-(f): Mathematical
simulation curves for different samples.
Fig.6(c) was a full-scan curve of Cu2+ solution with different concentrations after adding coloring
agent, and it can be seen from the figure that the optimal detection wavelength of Cu2+ was 448 nm. At
448 nm, Cu2+ solution absorbance showed a good linear relationship with the solution concentration. In
Fig.6(a,b), GL/CS-FA and GL/CS reached the adsorption balance in 10 h, and GL/CS-FA adsorption
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effect was worse than that of GL/CS. CS macromolecule played a role of adsorption Cu2+, and the poor
adsorption performance of GL/CS-FA indicated that FA occupied a part of the binding site of CS. The
fixation process GL/CS/MAP-FA microspheres is more complex, adsorption and precipitation conversion
are the two main fixation ways. When Cu2+ diffused into the microspheres, a small amount of Cu2+ was
adsorbed and fixed by CS. Most of the Cu2+ reacted with MAP to form insoluble copper phosphate, and
simultaneously released Mg2+ and NH4+. GL/CS/MAP-FA-1 owned a high GL content and exhibited a
rapid swelling characteristic. When the swelling was stable, MAP on the surface of the microspheres was
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basically consumed, and a short period of stabilization occurred in the fixing process. After a period of
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time, MAP inside the microspheres gradually began to react with Cu2+. The adsorption data were
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subjected to curve regression analysis, and the logarithmic curve was closer to the measured value of the
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adsorption data. The difference in sample composition led to differences in the R2 value of the logarithmic
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estimation curves. GL/CS/MAP-FA-1, 2, 3 (R2> 0.95) were more suitable for logarithmic function model
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Table 3. Parameters of Cu2+ removal kinetic analysis of GL/CS/MAP-FA microspheres according to different models.
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k1 R2 k2 R2 kint R2 C
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In order to study Cu2+ removal behavior of microspheres, intraparticle diffusion (Fig.6(d)), pseudo-
first order (Fig.6(e)) and pseudo-second order (Fig.6(f)) mathematical models were used [32]. The
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principle of the pseudo-first order model is the membrane diffusion theory. The limiting step of the
pseudo-second order model is the adsorption rate, and electron transfer and electron sharing phenomena
exist between the adsorbent and the adsorbate. The intra-particle diffusion model is suitable for predicting
the control step of intra-particle diffusion [33-35]. Equation (5)-(7) were applied to describe the Cu2+
removal process, and some important parameters were calculated (Table 3). The higher the model fits, the
closer the correlation coefficient (R2) is to 1. It can be seen that GL/CS/MAP-FA-2,3 conformed to
pseudo-second order model (R2>0.992). In addition, the fixation process of GL/CS/MAP-FA-1 was
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divided into two segments, and each segment was more suitable for pseudo-first order model (R2 >
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0.985). However, the fixation effect of GL/CS-FA and GL/CS was poor, and the difference of R2
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calculated by each model was small, which may be due to the Cu2+ adsorption-desorption process.
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3.7 Degradation and biological toxicity performance of GL/CS/MAP-FA microspheres
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The degradation experiments were performed in an aqueous environment [36]. The degradation column
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chart of GL/CS/MAP-FA microspheres together with the surface morphology under neutral and acidic
conditions was given in Fig.7. Under neutral conditions (Fig.7(a)), the degradation process of the
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microspheres was divided into three stages. The degradation rate was faster in the first 20 days, which
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was caused by the dissolution of FA and uncrosslinked macromolecules in water. About 20 days, the
dissolved substances in the microspheres basically diffused into the water, and the mass of the
microspheres remained stable. After 45 days, the degradation rate of the microspheres increased suddenly,
which may be caused by macromolecular hydrolysis or destruction of the crosslinking points in the three-
dimensional networks. Under acidic conditions (Fig.7(b)), the degradation process of the microspheres
was divided into two stages. First, the water-soluble substance diffused into the water. Then, the
hydrolysis and degradation of the macromolecules may be caused by H+ in the solution, and MAP may be
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slowly converted into soluble salts. Under above two conditions, the degradation rate of GL/CS/MAP-
FA-3 was relatively lower, and this may be ascribed to more content of CS in the microspheres. Nie et al.
revealed that high moisture content in the environment makes against microbes growing and inhibits the
degradation process [37]. The weightlessness data were subjected to curve regression analysis, and all
function estimation curves (R2 < 0.90) cannot simulate the measured values well. This was related to
The surface morphology of the microspheres, GL/CS/MAP-FA-1, degraded in different stages was shown
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in Fig.7(c). At first, the surface of the microspheres was even and only some small powders adhered. At
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45 days, the microspheres were severely eroded, and at 90 days, the microspheres were completely
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broken. The destruction of crosslinking points in the three-dimensional network of drug-loaded
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microspheres led to further release of fertilizer. Therefore, during plant growth, as the microspheres were
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degraded, most of the nutrients in the microspheres were released into the soil, which was beneficial to
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the growth of the plants. In addition, the materials for preparing the microspheres were made into a
culture medium and cultured at 37°C. After a period of time, the culture medium was covered with black
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colonies (Fig.7(d)). This showed that the preparation materials of the microspheres were non-toxic to the
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Fig.7 (a), (b): Degradation column chart of GL/CS/MAP-FA-1, 2, 3 microspheres under neutral and acidic conditions;
(c): SEM images of GL/CS/MAP-FA-1 microspheres degraded in water. (d): Colony growth process.
Fig.8 supplied the garlic growth column chart together with some photos of garlic, wheat, chrysanthemum
coronarium and corn in growth cycle. As can be seen in Fig.8(a), the plant height of the garlic in the blank
group was lower compared to the experimental groups, and its difference was continued to increase. An
approximate garlic growth curve can be obtained by smoothly connecting the vertices of the column. The
curves of garlic growth rate can be obtained by differentiating the garlic growth curves. From Fig.8(b), plants
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that added with microspheres grew significantly faster than that of the blank group.
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Fig.8 (a): Garlic growth column chart (microspheres: 0.2 g); (b): Garlic growth rate graph;
(c): Pictures of garlic, wheat, chrysanthemum coronarium and corn (microspheres: 0.2g; B0: blank; S1:
GL/CS/MAP-FA-1; S2: GL/CS/MAP-FA-2; S3: GL/CS/MAP-FA-3).
The N and P elements with the ion state in the soil are easy to run off, while the N and P elements of
the organic state are highly utilized [38, 39]. GL/CS/MAP-FA microspheres can be degraded into small
nitrogen-containing organic molecules. These molecules are beneficial to the survival of microorganisms
in the soil and the fertility of the land can be improved [40]. FA can reduce fertilizer loss, increase plant
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drought resistance, and improve nutrient absorption. The garlic added with GL/CS/MAP-FA microspheres
not only grew faster, but also possessed a relatively thicker trunk and bigger roots. Wheat,
chrysanthemum coronarium and corn were planted under the same conditions. From Fig.8(c), the plants
added with the GL/CS/MAP-FA microspheres were superior to the blank group in terms of plant height,
trunk diameter and root thickness. This was the same as the results of the garlic planting experiment.
These results showed that GL/CS/MAP-FA microspheres owned a promoting effect on the growth of
wheat, corn, garlic and other crops, and had no toxic effect on plants.
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4. Conclusions
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A novel type of GL/CS/MAP-FA soil conditioner microspheres was prepared using emulsion
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crosslinking method. The micro-morphology, sustained-release behavior, adsorption of heavy metal
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copper ions and degradation characteristics were studied. The sustained-release of GL/CS/MAP-FA
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microspheres conformed to the parabolic diffusion model, indicating that the slow-release of
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microspheres was controlled by the outer surface/edge diffusion process, which was accompanied with
ion exchange and dissociation process. The ion adsorption curves of heavy metal copper showed that the
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copper ion adsorption mechanism of microspheres was more complicated. The electron transfer and
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electron sharing phenomena existed between the adsorbed substance and the adsorbent, which followed
different adsorption kinetic models. Biological toxicity and degradation experiments revealed that the
microspheres had good biocompatibility, can be decomposed by microorganisms in the environment, and
In a word, GL/CS/MAP-FA microspheres owned good sustained-release and heavy metal adsorption
abilities and possessed good degradability. In addition, they were non-toxic, sustainable and
environmentally friendly, and can be used as a new type of soil conditioner in agricultural field.
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Acknowledgement
This work was funded by the projects of “Capacity Building Project of Some Local Colleges and
Technological Support Projects in the Field of Biomedicine (No. 19441901700)”, and “First-rate
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Discipline Construction of Applied Chemistry (No. 2018xk-B-06)”.
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