Biosynthesis and characterization of deuterated chitosan in filamentous fungus and yeast

Yue Yuan; Hui Li; Wellington Leite; Qiu Zhang; Peter V. Bonnesen; Jessy L. Labbé; Kevin L. Weiss; Sai Venkatesh Pingali; Kunlun Hong; Volker S. Urban; Sonja Salmon; Hugh O'Neill

doi:10.1016/j.carbpol.2021.117637

Biosynthesis and characterization of deuterated chitosan in filamentous fungus and yeast

Yue Yuan, Hui Li, Wellington Leite, Qiu Zhang, Peter V. Bonnesen, Jessy L. Labbé, Kevin L. Weiss, Sai Venkatesh Pingali, Kunlun Hong, Volker S. Urban, Sonja Salmon, Hugh O'Neill

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Abstract

Deuterated chitosan was produced from the filamentous fungus Rhizopus oryzae, cultivated with deuterated glucose in H₂O medium, without the need for conventional chemical deacetylation. After extraction and purification, the chemical composition and structure were determined by Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR), and small-angle neutron scattering (SANS). ¹³C NMR experiments provided additional information about the position of the deuterons in the glucoseamine backbone. The NMR spectra indicated that the deuterium incorporation at the non-exchangeable hydrogen positions of the aminoglucopyranosyl ring in the C3 – C5 positions was at least 60–80 %. However, the C2 position was deuterated at a much lower level (6%). Also, SANS showed that the structure of deuterated chitosan was very similar compared to the non-deuterated counterpart. The most abundant radii of the protiated and deuterated chitosan fibers were 54 Å and 60 Å, respectively, but there is a broader distribution of fiber radii in the protiated chitosan sample. The highly deuterated, soluble fungal chitosan described here can be used as a model material for studying chitosan-enzyme complexes for future neutron scattering studies. Because the physical behavior of non-deuterated fungal chitosan mimicked that of shrimp shell chitosan, the methods presented here represent a new approach to producing a high quality deuterated non-animal-derived aminopolysaccharide for studying the structure-function association of biocomposite materials in drug delivery, tissue engineering and other bioactive chitosan-based composites.

Original language	English
Article number	117637
Journal	Carbohydrate Polymers
Volume	257
DOIs	https://doi.org/10.1016/j.carbpol.2021.117637
State	Published - Apr 1 2021

Funding

This manuscript has been authored in part by UT-Battelle, LLC, under contract DE-AC05-00OR22725 with the US Department of Energy (DOE). The US government retains and the publisher, by accepting the article for publication, acknowledges that the US government retains a nonexclusive, paid-up, irrevocable, worldwide license to publish or reproduce the published form of this manuscript, or allow others to do so, for US government purposes. DOE will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan ( http://energy.gov/downloads/doe-public-access-plan ). This research was supported in part by an appointment for Y.Y. to the Nuclear Engineering and Science Laboratory Synthesis (NESLS) Program at Oak Ridge National Laboratory . The enzyme immobilization work inspiring this chitosan deuteration study is part of on-going research at North Carolina State University. Authors gratefully acknowledge the help of Dr. Flora Meilleur in discussing the structural characterization methods for immobilized enzymes. Authors acknowledge Tosoh Biosciences, LLC B io, Inc, for chromatography data collection. The SANS studies on Bio-SANS and biodeuteration were supported by the U.S. Department of Energy (DOE) Office of Biological and Environmental Research (OBER) funded Center for Structural Molecular Biology (CSMB) under Contract FWP ERKP291, using the High Flux Isotope Reactor supported by the Basic Energy Sciences , DOE. NMR characterization was conducted at the Center for Nanophase Materials Sciences, which is a DOE Office of Science User Facility. J.L. was supported by the Center for Bioenergy Innovation (CBI), a DOE OBER Bioenergy Research Center . This manuscript has been coauthored by UT-Battelle, LLC under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States government retains a non-exclusive, paid-up, irrevocable, world-wide license to publish or reproduce the published form of this manuscript, or allow others to do so, for United States government purposes. The Department of Energy will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan ( http://energy.gov/downloads/doe-public-access-plan ). ORNL is managed by UT-Battelle, LLC under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy.

Keywords

Biodeuteration
Enzyme entrapment
Microbial chitosan
NMR
Neutron scattering

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10.1016/j.carbpol.2021.117637

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title = "Biosynthesis and characterization of deuterated chitosan in filamentous fungus and yeast",

abstract = "Deuterated chitosan was produced from the filamentous fungus Rhizopus oryzae, cultivated with deuterated glucose in H2O medium, without the need for conventional chemical deacetylation. After extraction and purification, the chemical composition and structure were determined by Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR), and small-angle neutron scattering (SANS). 13C NMR experiments provided additional information about the position of the deuterons in the glucoseamine backbone. The NMR spectra indicated that the deuterium incorporation at the non-exchangeable hydrogen positions of the aminoglucopyranosyl ring in the C3 – C5 positions was at least 60–80 %. However, the C2 position was deuterated at a much lower level (6%). Also, SANS showed that the structure of deuterated chitosan was very similar compared to the non-deuterated counterpart. The most abundant radii of the protiated and deuterated chitosan fibers were 54 {\AA} and 60 {\AA}, respectively, but there is a broader distribution of fiber radii in the protiated chitosan sample. The highly deuterated, soluble fungal chitosan described here can be used as a model material for studying chitosan-enzyme complexes for future neutron scattering studies. Because the physical behavior of non-deuterated fungal chitosan mimicked that of shrimp shell chitosan, the methods presented here represent a new approach to producing a high quality deuterated non-animal-derived aminopolysaccharide for studying the structure-function association of biocomposite materials in drug delivery, tissue engineering and other bioactive chitosan-based composites.",

keywords = "Biodeuteration, Enzyme entrapment, Microbial chitosan, NMR, Neutron scattering",

author = "Yue Yuan and Hui Li and Wellington Leite and Qiu Zhang and Bonnesen, {Peter V.} and Labb{\'e}, {Jessy L.} and Weiss, {Kevin L.} and Pingali, {Sai Venkatesh} and Kunlun Hong and Urban, {Volker S.} and Sonja Salmon and Hugh O'Neill",

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doi = "10.1016/j.carbpol.2021.117637",

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T1 - Biosynthesis and characterization of deuterated chitosan in filamentous fungus and yeast

AU - Yuan, Yue

AU - Li, Hui

AU - Leite, Wellington

AU - Zhang, Qiu

AU - Bonnesen, Peter V.

AU - Labbé, Jessy L.

AU - Weiss, Kevin L.

AU - Pingali, Sai Venkatesh

AU - Hong, Kunlun

AU - Urban, Volker S.

AU - Salmon, Sonja

AU - O'Neill, Hugh

PY - 2021/4/1

Y1 - 2021/4/1

N2 - Deuterated chitosan was produced from the filamentous fungus Rhizopus oryzae, cultivated with deuterated glucose in H2O medium, without the need for conventional chemical deacetylation. After extraction and purification, the chemical composition and structure were determined by Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR), and small-angle neutron scattering (SANS). 13C NMR experiments provided additional information about the position of the deuterons in the glucoseamine backbone. The NMR spectra indicated that the deuterium incorporation at the non-exchangeable hydrogen positions of the aminoglucopyranosyl ring in the C3 – C5 positions was at least 60–80 %. However, the C2 position was deuterated at a much lower level (6%). Also, SANS showed that the structure of deuterated chitosan was very similar compared to the non-deuterated counterpart. The most abundant radii of the protiated and deuterated chitosan fibers were 54 Å and 60 Å, respectively, but there is a broader distribution of fiber radii in the protiated chitosan sample. The highly deuterated, soluble fungal chitosan described here can be used as a model material for studying chitosan-enzyme complexes for future neutron scattering studies. Because the physical behavior of non-deuterated fungal chitosan mimicked that of shrimp shell chitosan, the methods presented here represent a new approach to producing a high quality deuterated non-animal-derived aminopolysaccharide for studying the structure-function association of biocomposite materials in drug delivery, tissue engineering and other bioactive chitosan-based composites.

AB - Deuterated chitosan was produced from the filamentous fungus Rhizopus oryzae, cultivated with deuterated glucose in H2O medium, without the need for conventional chemical deacetylation. After extraction and purification, the chemical composition and structure were determined by Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR), and small-angle neutron scattering (SANS). 13C NMR experiments provided additional information about the position of the deuterons in the glucoseamine backbone. The NMR spectra indicated that the deuterium incorporation at the non-exchangeable hydrogen positions of the aminoglucopyranosyl ring in the C3 – C5 positions was at least 60–80 %. However, the C2 position was deuterated at a much lower level (6%). Also, SANS showed that the structure of deuterated chitosan was very similar compared to the non-deuterated counterpart. The most abundant radii of the protiated and deuterated chitosan fibers were 54 Å and 60 Å, respectively, but there is a broader distribution of fiber radii in the protiated chitosan sample. The highly deuterated, soluble fungal chitosan described here can be used as a model material for studying chitosan-enzyme complexes for future neutron scattering studies. Because the physical behavior of non-deuterated fungal chitosan mimicked that of shrimp shell chitosan, the methods presented here represent a new approach to producing a high quality deuterated non-animal-derived aminopolysaccharide for studying the structure-function association of biocomposite materials in drug delivery, tissue engineering and other bioactive chitosan-based composites.

KW - Biodeuteration

KW - Enzyme entrapment

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KW - NMR

KW - Neutron scattering

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VL - 257

JO - Carbohydrate Polymers

JF - Carbohydrate Polymers

M1 - 117637

ER -

Biosynthesis and characterization of deuterated chitosan in filamentous fungus and yeast

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