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Short Note

4-(Hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-D-mannofuranose

1
Ingénierie des Matériaux Polymères (IMP, UMR CNRS 5223), Université Claude Bernard Lyon 1, Université de Lyon, Villeurbanne F-69622, France
2
Department of Chemistry, Institut Teknologi Bandung, Bandung 40132, Indonesia
*
Author to whom correspondence should be addressed.
Molbank 2014, 2014(1), M815; https://doi.org/10.3390/M815
Submission received: 17 January 2014 / Accepted: 8 February 2014 / Published: 12 February 2014

Abstract

:
Low molecular weight chitooligosaccharide with one 2,5-anhydro-d-mannofuranose unit at the reducing end (COSamf) was prepared by nitrous deamination of fully N-deacetylated chitosan. The functionalization of the amf unit by reductive amination with 4-(hexyloxy)aniline in presence of NaBH3CN was achieved in high yield. The chemical structure of the targeted 4-(hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-d-mannofuranose was fully characterized by NMR spectroscopy, MALDI-TOF mass spectrometry and size-exclusion chromatography. This synthesis opens the way to a new generation of COSamf derivatives with potential amphiphilic properties.

Graphical Abstract

Chitosan is a random linear polysaccharide of d-glucosamine (GlcN) and N-acetyl-d-glucosamine (GlcNAc) units linked by β-(1→4) glycosidic bonds. Chitosan is generally obtained by chemical or enzymatic N-deacetylation of chitin, the second most abundant naturally occurring polymer produced industrially from shells of crustaceans and squid pens [1,2,3,4]. Chitooligosaccharides (COS), also named chitosan or chitin oligomers, have recently received considerable attention as functional biomolecules with a wide range of applications in food, agriculture, medicine, pharmaceutics and cosmetics. COS take advantage of their various interesting physico-chemical and biological properties, including water-solubility, biocompatibility, antibacterial, antifungal and antitumoral activities [5,6,7,8,9]. In order to improve the scope of their properties, chemical modifications of COS have been investigated for a decade [10,11,12]. In this study, we described the synthesis of the 4-(hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-d-mannofuranose. The interest of this work is to take advantage of the reactivity of the aldehyde group of the 2,5-anhydro-d-mannofuranose (amf) unit present at the reducing end of COS obtained by nitrous deamination of chitosan, to generate original amphiphilic COS derivatives.

Results and Discussion

4-(hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-d-mannofuranose was efficiently synthesized from chitosan in a two-step procedure involving the reductive amination of chitooligosaccharide-2,5-anhydro-d-mannofuranose (COSamf, 1) with 4-(hexyloxy)aniline as illustrated in Scheme 1.
COSamf 1 was prepared by nitrous acid deamination of a fully N-deacetylated chitosan based on the method previously described by Tommeraas et al. [13]. Thus, the depolymerization of chitosan (DA 0%, Mw = 270 kg/mol; Mn = 115 kg/mol, Ð = 2.3) by NaNO2 (GlcN/NaNO2 molar ratio = 10) in aqueous acid solution at room temperature led to COSamf 1 in 67% mass yield after 24 h of reaction. The chemical structure of COSamf 1 was fully confirmed by 1H and 13C-NMR spectroscopies, MALDI-TOF mass spectrometry and size-exclusion chromatography (see Supporting Information). Therefore it has been shown COSamf 1 is composed of a mixture of oligomers, with an average number of GlcN units into chains around 23.
The reductive amination of COSamf 1 with 4-(hexyloxy)aniline in presence of NaBH3CN was carried out at 40 °C in buffer solution (pH 5.5) for 48 h, leading to the targeted 4-(hexyloxy)aniline-linked COSamf 2 in an excellent mass yield (92%). The chemical structure of the title compound was entirely characterized by 1H and 13C-NMR spectroscopies thanks to two-dimensional NMR analyses, pointing out the coupling reaction between the aldehyde function of COSamf 1 and the amine group of the aniline residue. Thus, the presence of the corresponding CH2-N covalent linkage was displayed at δ 3.50 ppm for methylene protons and 69.7 ppm for the methylene carbon, respectively in 1H and 13C-NMR spectra. As confirmed by MALDI-TOF mass spectrometry (see Supporting Information), 4-(hexyloxy)aniline-linked COSamf 2 is composed of a mixture of oligomers, with an average number of GlcN units into chains, determined by both 1H-NMR and SEC, equal to 23 as for COSamf 1.

Experimental

4-(Hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-d-mannofuranose 2: A fully N-deacetylated chitosan (2.1 g, 13 mmol GlcN unit) was solubilized in 1 L of water by addition of 11.5 mL HCl (37% w/w). A freshly prepared solution of NaNO2 (1.3 mmol) was added and the reaction was allowed to proceed for 24 h at room temperature. The product was precipitated by addition of conc. NH4OH, centrifuged (15 min, 11200 rpm), washed with distilled water until neutral pH, then freeze-dried leading to COSamf 1 (1.4 g, 67% mass yield) as a white powder. COSamf 1 (0.5 g, 0.14 mmol of amf unit) was then solubilized in 20 mL of ammonium acetate buffer (50 mM, pH 5.5). 271 mg of 4-(hexyloxy)aniline (1.4 mmol) in 10 mL ethanol and 88 mg of sodium cyanoborohydride (1.4 mmol) were added and the reaction was allowed to proceed for 48 h at 40 °C. The product was precipitated by addition of conc. NH4OH, centrifuged (15 min, 11,200 rpm), washed with water/ethanol (50:50) then freeze-dried leading to 2 (460 mg, 92% mass yield) as a white powder. 1H-NMR (300 MHz, D2O, 298 °K): δ (ppm) 7.45 (d, J = 9.0 Hz, 2H, H aromatic), 7.15 (d, J = 9.0 Hz, 2H, H aromatic), 4.90–4.70 (m, 23H, H-1 GlcN), 4.32 (m, 1H, H-3 amf), 4.24 (m, 1H, H-5 amf), 4.18 (m, 1H, H-4 amf), 4.14 (m, 1H, H-2 amf), 4.08 (t, J = 6.6 Hz, 2H, CH2O), 4.00–3.40 (m, H-3 to H-6 GlcN, H-6 amf, CH2N), 3.18 (t, J = 8.9 Hz, 23H, H-2 GlcN), 1.75 (m, 2H, CH2), 1,40 (m, 2H, CH2), 1.30 (m, 4H, 2CH2), 0.85 (t, J = 7.0 Hz, 3H, CH3). 13C-NMR (125 MHz, D2O, 298 °K): δ (ppm) 159.8 (CO aromatic), 127.3 (CN aromatic), 124.7 (2CH aromatic), 116.8 (2CH aromatic), 99.3 (C-1' GlcN), 98.1 (C-1 GlcN), 86.8 (C-4 amf), 83.1 (C-5 amf), 78.6 (C-2 amf), 77.9 (C-3 amf), 77.0 (C-5’ GlcN), 76.9 (C-4 GlcN), 75.3 (C-5 GlcN), 72.3 (C-3' GlcN), 70.6 (C-3 GlcN), 70.2 (C-4' GlcN), 69.7 (CH2O), 61.9 (C-6 amf), 60.9 (C-6' GlcN), 60.6 (C-6 GlcN), 56.4 (C-2 GlcN), 56.1 (C-2' GlcN), 53.2 (CH2N), 31.3 (CH2), 28.7 (CH2), 25.4 (CH2), 22.5 (CH2), 13.9 (CH2). Note that C’ represents carbon atoms of the GlcN unit linked to the amf unit. MALDI-TOF MS: presence of a major peak at m/z 1650.5 attributed to HO-(GlcN)8-C18H28NO4 (m/z monoisotopic calcd for [C66H117O37N9Na]+ = 1650.7 mass units (Δ = 0.01%)). HRMS (ESI): calcd for C66H117O37N9Na: m/z 1650.7448; found 1650.7432 [M+Na]+ (difference = 1.6 ppm).

Supplementary materials

Supplementary File 1Supplementary File 2Supplementary File 3Supplementary File 4

Acknowledgments

The authors thank Catherine Ladavière (IMP, CNRS), Agnès Crepet (IMP, CNRS) and Bernard Fenet (CCRMN, Université Lyon 1) for their helpful assistances and discussions in MALDI-TOF mass spectrometry, size-exclusion chromatography and NMR spectroscopy analyses, respectively. ES specially thanks Didin Mujahidin (Organic Chemistry Laboratory of Institut Teknologi Bandung, Indonesia) for his fruitful support during this study. Financial support from the University Lyon 1 is greatly acknowledged.

Conflicts of Interest

The authors declare no conflict of interest.

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Scheme 1. Synthesis of the 4-(hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-d-mannofuranose from chitosan.
Scheme 1. Synthesis of the 4-(hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-d-mannofuranose from chitosan.
Molbank 2014 m815 sch001

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MDPI and ACS Style

Salim, E.; Galais, A.; Trombotto, S. 4-(Hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-D-mannofuranose. Molbank 2014, 2014, M815. https://doi.org/10.3390/M815

AMA Style

Salim E, Galais A, Trombotto S. 4-(Hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-D-mannofuranose. Molbank. 2014; 2014(1):M815. https://doi.org/10.3390/M815

Chicago/Turabian Style

Salim, Emil, Alice Galais, and Stéphane Trombotto. 2014. "4-(Hexyloxy)aniline-linked chitooligosaccharide-2,5-anhydro-D-mannofuranose" Molbank 2014, no. 1: M815. https://doi.org/10.3390/M815

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