Patent Application: US-200913054314-A

Abstract:
α - lactosamine hydrochloride , substantially free of β - lactosamine hydrochloride ; a method of preparing α - lactosamine hydrochloride monohydrate from an anomeric mixture of lactosamine hydrochloride , including : preparing a solution including the anomeric mixture of lactosamine hydrochloride , water and at least one water - miscible organic solvent at a temperature of 0 - 100 ° c ., and cooling the solution to cause crystallization of α - lactosamine hydrochloride monohydrate ; a method of preparing anhydrous α - lactosamine hydrochloride ; and use of α - lactosamine hydrochloride as a food supplement or intermediate in synthesis .

Description:
a crude anomeric mixture of lactosamine hydrochloride can be prepared by using literature methods . 1 - 7 different methods of preparation provide different purities of the crude mixture . however , all the expected impurities and side products present in the reaction mixtures obtained by the different methods are found to dissolve in aqueous alcohols at 40 - 100 ° c . temperatures . stirring a solution of the crude mixture in a solvent or mixture of solvents as set out above at between − 5 and 30 ° c . initiates the crystallization of α - lactosamine hydrochloride . filtration , washing of the crystals with etoh or iso - propanol , followed by a drying process at room temperature or higher temperatures provides high purity α - lactosamine hydrochloride monohydrate with a melting point of 174 ° c . alternatively , the crude lactosamine hydrochloride can be dissolved in water at 40 - 100 ° c . and subsequently the at least one water miscible solvent is added . crystallization of α - lactosamine hydrochloride starts upon cooling and stirring . 1 . 5 kg crude anomeric mixture of lactosamine hydrochloride obtained according to literature procedures &# 39 ; was dissolved in warm water ( 1 . 5 l , 50 ° c .). ethanol ( 4 l ) was added to the mixture over a period of 30 min . then the mixture was cooled down to 0 ° c . and stirred for 8 h . the white crystals were collected by filtration and washed with ethanol ( 200 ml ). yield : 1 . 3 kg pure alpha anomer . 100 g crude anomeric mixture of lactosamine hydrochloride obtained according to literature procedures 7 was dissolved in warm water ( 100 ml , 50 ° c .). i - propanol ( 250 ml ) was added to the mixture over a period of 10 min . then the mixture was cooled down to 0 ° c . and stirred for 8 h . the white crystals were collected by filtration and washed with i - propanol ( 20 ml ). yield : 85 g pure alpha anomer . 100 g crude anomeric mixture of lactosamine hydrochloride obtained according to literature procedures 7 was dissolved in warm water ( 100 ml , 50 ° c .). the solution was added to acetone ( 1 l ) obtaining a white precipitate . this powder was redissolved in water ( 90 ml ) and i - propanol ( 220 ml ) added to the mixture over a period of 10 min . then the mixture was cooled down to 0 ° c . and stirred for 8 h . the white crystals were collected by filtration and washed with i - propanol ( 20 ml ). yield : 82 g pure alpha anomer . 100 g crude anomeric mixture of lactosamine hydrochloride obtained according to literature procedures 7 was dissolved in warm water ( 100 ml , 50 ° c .). the solution added to acetone ( 1 l ) obtaining a white precipitate . this powder was redissolved in water ( 90 ml ) and ethanol ( 300 ml ) added to the mixture over a period of 10 min . then the mixture was cooled down to 0 ° c . and stirred for 8 h . the white crystals were collected by filtration and washed with ethanol ( 20 ml ). yield : 78 g pure alpha anomer . 100 g crude anomeric mixture of lactosamine hydrochloride obtained according to literature procedures 7 was dissolved in warm water ( 100 ml , 50 ° c .). the solution was added to acetone ( 1 l ) obtaining a white precipitate . this powder was redissolved in water ( 90 ml ) and a mixture of ethanol and i - propanol ( 8 % i - propanol , 220 ml ) was added to the mixture over a period of 10 min . then the mixture was cooled down to 0 ° c . and stirred for 8 h . the white crystals were collected by filtration and washed with the solvent mixture ( 20 ml ). yield : 80 g pure alpha anomer . 100 g crude anomeric mixture of lactosamine hydrochloride obtained according to literature procedures &# 39 ; was dissolved in warm water ( 100 ml , 50 ° c .). a mixture of ethanol and i - propanol ( 8 % i - propanol , 250 ml ) was added to the mixture over a period of 10 min . then the mixture was cooled down to 0 ° c . and stirred for 8 h . the white crystals were collected by filtration and washed with the solvent mixture ( 20 ml ). yield : 88 g pure alpha anomer . 100 g alpha lactosamine hydrochloride obtained according to procedures above was dehydrated under high - vacuum ( 1 - 10 mbar ) at the temperature of 55 ° c . resulting in 95 g dehydrated pure alpha anomer . 100 g alpha lactosamine hydrochloride obtained according to procedures above was dehydrated under high - vacuum ( 1 - 10 mbar ) at the temperature of 75 ° c . resulting in 95 g dehydrated pure alpha anomer . 100 g alpha lactosamine hydrochloride obtained according to procedures above was dehydrated under high - vacuum ( 10 - 15 mbar ) at the temperature of 40 ° c . resulting in 95 g dehydrated pure alpha anomer . 100 g alpha lactosamine hydrochloride obtained according to procedures above was dehydrated under high - vacuum ( 10 - 15 mbar ) at the temperature of 75 ° c . resulting in 95 g dehydrated pure alpha anomer . 100 g alpha lactosamine hydrochloride obtained according to procedures above was dehydrated under high - vacuum ( 20 - 50 mbar ) at the temperature of 95 ° c . resulting in 94 g dehydrated pure alpha anomer . 100 g alpha lactosamine hydrochloride obtained according to procedures above was dehydrated under high - vacuum ( 50 - 100 mbar ) at the temperature of 120 ° c . resulting in 94 g dehydrated pure alpha anomer . lactosamine hydrochloride sample was made according the present invention for single crystal x - ray diffraction studies . no crystal structure of lactosamine hydrochloride was published before . however , x - ray structure of the n - acetyllactosamine is known . single crystals suitable for x - ray diffraction measurement could be grown by layering methanol onto water solution of lactosamine hydrochloride . this is carried out by preparing a saturated aqueous solution of lactosamine hydrochloride , filtering the solution to remove any remaining solid lactosamine hydrochloride , and then adding a small volume of water followed by careful addition of methanol . a rather small colorless prism crystal was chosen for the measurement and fixed on the loop using perfluorinated paraffin oil . structural data were reliable and the results are acceptable for scientific publication and / or patenting . colorless prism crystal ( 0 . 08 × 0 . 05 × 0 . 02 mm ) of c 12 h 26 n 1 cl 1 o 11 , m = 395 . 79 , monoclinic crystal system , a = 4 . 7931 ( 8 ) å , b = 13 . 5508 ( 19 ) å , c = 13 . 2816 ( 19 ) å , β = 93 . 97 ( 1 )°, v = 860 . 6 ( 2 ) å 3 , z = 2 , space group : p2 1 , ρ calc = 1 . 527 g cm − 3 . data were collected at 110 ( 1 ) k , bruker - gadds multiwire proportional diffractometer , cu kα radiation λ = 1 . 54184 å , θ max = 63 . 3 °, 6882 measured , 2059 independent reflections of which 1793 reflections were unique with i & gt ; 2σ ( i ). raw data was evaluated using the frambo software , the structure was solved using the sir - 92 software and refined on f 2 using shelx - 97 program , publication material was prepared with the wingx - 97 suite , r ( f )= 0 . 061 and wr ( f 2 )= 0 . 156 for 2509 reflections , 254 parameters , 10 restraints . residual electron density : 0 . 33 /− 0 . 33 e / å 3 . hydrogen atoms at carbon atoms were placed into geometric position while other hydrogen atoms could be found at the difference electron density map . distance of these hydrogen atoms to the respective oxygen or nitrogen was constrained . coordinates of hydrogen atoms were allowed to refine within a short range to the donor atom . position of the hydrogens on the amino group was found using the riding model and their distance to the nitrogen atom was fixed . because of the extensive hydrogen bond network , alternative positions for some of these hydrogen atoms are possible . full data and the structure determined therefrom are shown in fig1 and 2 . 10 mg pure α - lactosamine hydrochloride was dissolved in 700 μl d 2 o , then 1h - nmr spectrum was recorded immediately . a second 1h - nmr spectra was recorded a day later , by which time the clean substance had already anomerized . on first spectrum ( fig3 , above ) at 5 . 26 ppm the h - 1 and at 4 . 29 ppm the h - 1 ′ can be seen . on the second spectrum ( fig3 , below ) next to the alpha anomer already the beta appears . at 4 . 78 ppm h - 1 - beta and at 2 . 85 ppm h - 2 - beta can be seen . this experiment shows that the starting material is a clean anomer , which anomerizes at room temperature within one day . 1 . oligosaccharides as antitumor drugs and health foods . misawa , yoshitomo ; 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