Chitin from shrimp shells (practical grade coarse flakes), chitosan CHIT50.1 (Product number: 900,344) and CHIT50.2 (Product number: 448,869), N-acetyl-D-glucosamine (GlcNAc) and biotin were from Sigma Aldrich (St. Louis, MO, USA). Chitosan from shrimp shells CHIT100 and CHIT600 were from Across Organics Chemicals (ThermoFisher Scientific, Waltham, Massachusetts, USA). Colloidal chitin (CC) was obtained as referred previously [30 (link)]. Basically, 10 g chitin in 10 M HCl was maintained 16 h at 4 ºC, filtered through glass thick fibres into ethanol and then chitin floccules were precipitated at 4 ºC, collected (5000xg during 10 min) and maintained in 0.1 M sodium acetate pH 4.0-4.5. Chitosan was dissolved in 0.1 M sodium acetate pH 4.0-4.5, in a final concentration of 10 g l− 1. Diacetyl-chitobiose ((GlcNAc)2), triacetyl-chitotriose ((GlcNAc)3), tetraacetyl-chitotetraose ((GlcNAc)4), pentaacetyl-chitopentaose ((GlcNAc)5) and hexaacetyl-chitohexaose ((GlcNAc)6) were from Megazyme (Bray, Irland). Nitrogen base w/o amino acids (YNB) was from Difco (BD, Sparks, MD, USA). Yeast extract, peptone, tryptone and agar were from Laboratorios Conda S.A. (Madrid, Spain).
Diacetyl chitobiose
Manufactured by Megazyme
Sourced in United States
Diacetyl-chitobiose is a disaccharide composed of two N-acetylglucosamine (GlcNAc) units. It is a substrate commonly used in assays and research applications involving enzymes that cleave glycosidic bonds, such as chitinases and β-N-acetylhexosaminidases.
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2 protocols using diacetyl chitobiose
1
Chitin and Chitosan Characterization
2
Cultivating Aeromonas salmonicida in Chitin
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Al. salmonicida strain LFI1238 originally isolated from the head kidney of diseased farmed cod (Gadhus morhua; (29) at 6000 × g for 1 minute, followed by immediate resuspension of the pellet in 1 mL Asmm.
The cell suspension was transferred to the final cultures by a 1:50 dilution in media supplemented with 0.2% glucose, 0.2% N-acetyl-D-glucosamine, 0.2 % diacetyl-chitobiose (Megazyme, Bray, County Wicklow, USA) or 1 % β-chitin from squid pen purchased from
France Chitine (Batch 20140101, Orange, France). Culture volumes ranged from 5-50 mL.
Final cultures were incubated at 12 °C with shaking at 175 rpm. Growth was measured by optical density (OD 600 ) using Ultrospec® 10 Cell Density Meter (Biochrom). The baseline was set by using sterile Asmm media with or without 1% β-chitin. OD 600 measurements of the β-chitin cultures was performed by allowing the cultures to settle for 30 seconds before collecting 1 mL for measurement.
The cell suspension was transferred to the final cultures by a 1:50 dilution in media supplemented with 0.2% glucose, 0.2% N-acetyl-D-glucosamine, 0.2 % diacetyl-chitobiose (Megazyme, Bray, County Wicklow, USA) or 1 % β-chitin from squid pen purchased from
France Chitine (Batch 20140101, Orange, France). Culture volumes ranged from 5-50 mL.
Final cultures were incubated at 12 °C with shaking at 175 rpm. Growth was measured by optical density (OD 600 ) using Ultrospec® 10 Cell Density Meter (Biochrom). The baseline was set by using sterile Asmm media with or without 1% β-chitin. OD 600 measurements of the β-chitin cultures was performed by allowing the cultures to settle for 30 seconds before collecting 1 mL for measurement.
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