CH630096A5 - Process for the preparation of nitroaromatic glycosides - Google Patents

Description

The invention relates to a process for the preparation of nitroaromatic glycosine of the formula (I)

ch2oh

(I),

in which n is an integer from 2, 3 or 4 and R is a substituted aromatic radical of the formula or in which X and Y independently of one another are H, no2, halogen, alkyl having 1 to 4 carbon atoms, OR1 or co2r1, with R1 is an alkyl radical with 1 to 6 carbon atoms with the proviso that at least one of the radicals X and Y is no2.

US Pat. No. 3,879,263 describes the determination of the a-amylase content of biological samples by adding maltotetraose or maltopentaose to a sample at constant temperature and constant pH. The method enables rapid determination of a-amylase and can be used to differentiate between a-amylase in saliva and a-amylase in pancreatic secretion. The latter forms glucose, while the former does not form glucose. The glucose formed can be spectrophotometric, e.g. can be determined by absorption of nicotinamide adenine dinu cluotide (reduced form) (NADH) at 340 nm. Since this method of determination depends on glucose, a glucose detection reaction is necessary. Furthermore, glucose present in the sample must either be removed or compensated for. The compounds according to the invention differ from this in that

as 4-nitrophenol is released as a substance which can then be related to a-amylase. This makes the determination independent of the step for the detection of glucose.

A.P. Jansen and P.G.A.B. Wydeveld found in Nature 162 (1958) 525 that a- (p-nitrophenyl) maltoside could be suitable as a substrate for the amylase determination. However, this publication shows that the authors never identified the active agent responsible for their observations. They report: 1) 4-nitrophenol was formed by incubating human urine or saliva samples with a- (p-nitrophenyl) maltoside at 37 ° C for 16 hours, which was identified 60 spectrophotometrically by mixing the hydrolyzate with 0.02N sodium hydroxide . 2) The hydrolysis was carried out by protein precipitants, e.g. 10% trichloroacetic acid and 0.5N silver nitrate prevented. 3) The hydrolysis was pH dependent and most effective at pH 5.9 65 to 7.0. The authors state that this was evidence of “the possible presence of an unidentified car-bohydrase”. It is not believed that a- (4-nitrophenyl) maltoside is suitable for an amylase determination

630096

is because the cleavage of this compound by a-amylase is extremely slow.

ch2oh

/

OH

oh ch-2oh

The process according to the invention for the preparation of nitroaromatic glycosine of the formula (I)

ch2oh

0

(I)

in which n is an integer from 2, 3 or 4 and R is a substituted aromatic radical of the formula or in which X and Y independently of one another are H, no2, halogen, alkyl having 1 to 4 carbon atoms, OR1 or co2r1, with R1 is an alkyl radical with 1 to 6 carbon atoms with the proviso that at least one of the radicals X and Y is no2,

is characterized in the preceding claim 1.

The use of the compounds produced according to the invention is in the preceding claim 10

25 characterized.

The nitroaromatic glycosides obtained according to the invention are descendants of a number of oligomers and polymers of glucose linked by a [- ^ 4]. This series of glycosides has the following general formula. The Gn nomenclature is useful for the short name of the a [l- »4] -linked glucose units.

HO

CHoOH oh cho0h ci oh ch2oh

"J4> -

Oh

0

CHpOH

Toilet n

oh a-D-glucose (Gi) n = 0 maltose (g2)

n = 1 maltotriose (G3) n = 2 maltotetraose (G4) n = 3 maltopentaose (G5) n = 4 maltohexaose (G6) n = 2000 amylose

Nomenclature of glucose [1-4] oligosaccharides

For the sake of brevity, the trial names and 45 short names given in Table I are used in the description of the invention. These refer to the systematic names given in Table I. The rules for this are set out in “Naming and Indexing of Chemical Substances for Chemical Abstracts Düring the Ninth Collective Period (1972-1976)”, Chemical Abstracts Service, according to Columbus, Ohio (1973).

Table I

Short name trivial name

Systematic designation

Gì g2 g3

g4

Gs

Ge

D-glucose

Maltose

Maltotriose

Maltotetraose

Maltopentaose

Maltohexaose

D-glucose

4-O-a-D-glucopyranosyl-D-glucose

0-a-D-glucopyranosyl- (1-4) -0-a-D-glucopyranosyl- (1-4) -D-glucose

0-a-D-glucopyranosyl- (1-4) -0-a-D-glucopyranosyl- (1-4) -0-a-D-glucopyranosyl- (1-4) -D-glucose

0-aD-glucopyranosyl- (1-4) -0-aD-glucopyranosyl- (1-4) -0-aD-glucopyranosyl- (1-4) -0-aD-glucopyranosyl- (1-4) -D -glucose

0-a-D-glucopyranosyl- (1-4) -0-a-D-glucopyranosyl- (1-4) -0-

a-D-glucopyranosyl- (l- * 4) -0-a-D-glucopyranosyl- (l- »4) -0-a-

D-glucopyranosyl- (1- * 4) -D-glucose

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Table I (continued) Nomenclature of glucose- [1 - * 4] oligosaccharides

Short name trivial name

Systematic designation

G4pNp a- (4-nitrophenyl) maltotetraoside

GsPnP a- (4-nitrophenyl) maltopentaoside

G3 (Ac) io deccaacetylmaltotriosyl G4 (Ac) i3 tridecaacetylmaltotetraosyl

4-nitrophenyl-0-aD-glucopyranosyl- (1-4) -0-aD-glucopyra-nosyl- (1 - * 4) -0-aD-glucopyranosyl- (1-4) -aD-glucopyranoside 4-nitrophenyl- 0-aD-glucopyranosyl- (1-4) -0-aD-glucopyra-nosyl- (1--4) -0-aD-glucopyranosyl- (1-4) -0-aD-glucopyrano-syl- (1st —4) -aD-glucopyranoside

2,3,6,2 ', 3', 6 ', 2' ', 3 ", 4", 6 "-decaacetyl-O-cc-D-glucopyranosyl- (1-4) -0-aD-glucopyranosyl- (1 - ^ 4) -0-aD-glucopyranosyl 2,3,6,2 ', 3', 6 ', 2 ", 3", 6 ", 2"', 3 "', 4"', 6 " '-Tridecaacetyl-OaD-glucopyranosyl- (1-4) -0-aD-glucopyranosyl- (1-4) -0-aD-glucopyranosyl- (1-4) -0-aD-glucopyranosyl

The glucose oligomers used as starting materials, i.e. Maltotetraose, maltopentaose and maltohexaose can be made by the process, either that described by R.L. Whistler and co-workers in J. Amer. Chem. Soc. 76 (1954) 1671 and 77 (1955) 1017,5761 or by Thomas John Pankratz in the applicant's US patent application 675,649. The preferred nitroaromatic glycosides of G4 and Gs can be prepared from pure G4 and Gs, which can be obtained by chromatography of the hydrolyzate of amylose as described by W. Pigman in "The

Carbohydrates », Academic Press, New York 1957, pages 678-679. 2o Reaction Scheme II illustrates a process for the preparation of the preferred compounds according to the invention. The details of each stage are described below.

The individual stages of the process for the preparation of the 2s compounds according to the invention are described in detail below.

CH20H

OH

Gn = G3 or G4

Ac20, NaOAc

"

CHz0Ac

Gn = G3 (Ac) i o or G4 (Ac) 13

CeHsOH, ZnCh

A

CHjOAc

CHJOäc

Gn = G3 (Ac) io or G4 (Ac) i3

oc6h5

Gn = G3 (Ac) io or G4 (Ac) i:

NoOMe

MeOH

CH20H

Gn = MALTO-N-OSIDE RESIDUE ct [1-4] Ac = CH3CO

n0z G3 (Ac) io = DECAACETYLMALTOTRIOSYL

G4 (Ac) i3 = TRIDECAACETYLMALTOTETRAOSYL

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Acetylation reaction

The acetylation of glucose oligomers with a mixture of acetic anhydride and anhydrous sodium acetate at elevated temperatures is known (WJ Whelan and PJ Roberts, J. Chem. Soc. (1953) 1298, WJ Whelan, JM Bailey and PJP Roberts, J. Chem. Soc. (1953) 1293, A. Thompson and ML Wolfrom, J. Amer. Chem. Soc. 74 (1954) 3612, ML Wolfrom, LW Georges, A. Thompson and IL Miller, J. Amer. Chem. Soc. 71 (1949) 2875. The fully acetylated derivatives with the substituent on the anomeric carbon atom are obtained in the β configuration, and the use of zinc chloride or acids as a catalyst instead of sodium acetate favors the configuration in this position (W. Pigman , loc. cit., pages 140-142), however, in the context of the invention the β configuration is desired so that the phenol in the next step can displace the anomeric acetoxy group by an inversion mechanism and form an a-phenylglycoside bond tion is carried out in acetic anhydride as solvent and reactant. The amount of acetic anhydride corresponds to 5 to 50 times the weight of G4 or Gs. 5 to 1 times the weight of G4 or Gs is preferred, so that sufficient reagent is present to keep the reactants in solution and to isolate the Allow product when the reaction mixture is poured into water. The anhydrous sodium acetate can be used in an amount of 1 to 10, preferably 5 to 6, molar equivalents per molar equivalent of G4 or Gs.

The reaction temperature can range from 100 ° C to 140 ° C, i.e. at the reflux temperature of acetic anhydride are preferably between 110 and 120 ° C. Below 100 ° C the acetylation is very slow and incomplete, and above 140 ° C (e.g. in a pressure vessel) the reaction is very violent and leads to a dark-colored product.

The reaction time can be 1 to 6 hours and is preferably about 2 hours. Prolonged heating at elevated temperatures also gives a dark product. The use of the reaction is noticeable in that the reaction mixture becomes a homogeneous solution and the reaction becomes slightly exothermic.

The completely acetylated product is isolated by pouring the cooled reaction mixture into ice water, the amount of which corresponds to 5 to 20 times the volume of the acetic anhydride, the mixture is stirred vigorously for a few minutes and then left to stand at 0 ° to 5 ° C. for at least 24 hours. The solid product, the crystallization of which can optionally be improved and accelerated by seeding, is filtered off, dried in air and recrystallized from a suitable solvent such as ethanol or methanol.

The identity of these and other intermediates is confirmed by the usual spectral properties and analyzes. The stereochemistry of the anomeric carbon can be easily determined by nuclear magnetic resonance spectroscopy of the protons ('H NMR) especially at high frequencies, e.g. Determine 220 MHz and if necessary by examining the optical rotation. In the NMR spectrum of aldopyranose acetates, an anomeric proton in the a configuration (Hia) can be distinguished from an anomeric proton in the β configuration (Hiss). In the NMR spectrum of aldopyranose acetates, an anomeric proton in the a configuration (Hia) has a chemical shift (8) near 5.75 ppm downfield from the inner tetramethysilane, and the signal appears as a doublet with an axial-axial coupling constant ( J) from

7 to 9 Hz. An anomeric proton in the β configuration (hot) gives a signal of 0.2 to 0.65 ppm downfield from this position and also has a doublet with an axially-equatorial or equatorial-equatorial coupling constant of 3 to 4 Hz (see, for example, LM Jackman “Applications of NMR Spectroscopy in Organic Chemistry”, pages 86 and 116, Pergamon Press (London 1959).

Acetate displacement reaction

The displacement of the anomeric acetate group in a fully acetylated sugar is easier than that of the other acetate groups. This is a useful property for synthetic purposes because it enables a preferred reaction at this position. For example, when tetradeca-acetylmaltotetraoside or heptadecaacetylmaltopentaoside is stirred with phenol and anhydrous zinc chloride (ZnCk) at elevated temperatures, the anomeric ß-acetoxy group is replaced by an a-phenoxy group. While at least one molar equivalent of phenol is required per molar equivalent of acetyl compound to meet the stoichiometry of the reaction, the reaction can be carried out with 3 to 20 molar equivalents, preferably 4 to 8 molar equivalents, of phenol to provide enough material to form one homogeneous solution is present. The amount of zinc chloride can be 0.25 to 5 molar equivalents per molar equivalent of acetyl compound and is preferably in the range of 0.5 to 1.5 molar equivalents. This reaction can be carried out at temperatures in the range from 80 ° C. to 120 ° C., temperatures in the range from 100 to 110 ° C. being preferred for the same reasons that were mentioned above for the first stage of the process. The reaction time can be 0.25 to 6 hours. Generally 1 to 3 hours will suffice. A solution of zinc chloride in a mixture of acetic acid and acetic anhydride (for example with a volume ratio of 95: 5) is an advantageous modification of the above-mentioned solvent and catalyst system for the introduction of the a-phenoxy group.

The anhydrous zinc chloride serving as a catalyst can be converted by acids, e.g. p-toluenesulfonic acid, and by other anhydrous covalent metal chlorides, e.g. Titanium (IV) chloride (TiCU), tin (IV) chloride (SnCk) and iron (III) chloride (FeCb). The displacement reaction is solvent-free because of the low melting point of the phenol (43 ° C) It is ensured that when excess phenol is used to bring the reaction to completion, the mixture remains a homogeneous solution at the reaction temperature. Similar reactions can also be expected if excess amounts of phenols that are liquid at the reaction temperature are used as both solvents and reactants, e.g. 2-cresol (melting point 30 ° C),

3-cresol (melting point 11 ° C), 4-cresol (melting point 35 ° C), 2-chlorophenol (melting point 8 ° C), 3-chlorophenol (melting point 29 ° C), 4-chlorophenol (melting point 37 ° C),

4-bromophenol (melting point 64 ° C), 2-nitrophenol (melting point 45 ° C), 2-methoxyphenol (guaiacol, melting point 32 ° C), 4-methoxyphenol (melting point 53 ° C), 2-methyl-5-isopropylphenol ( Carvacrol, melting point 1 ° C), 2-isopropyl-5-methylphenol (thymol, melting point 51 ° C) and methyl salicylate (melting point -8 ° C). For higher melting point phenols as well as the above phenols it is also possible to carry out the reaction in a solvent e.g. Benzene (boiling point 80 ° C), toluene (boiling point 110 ° C) or heptane (boiling point 98 ° C). When using the higher melting phenols, e.g. 3-nitrophenol (melting point 96 ° C), 4-nitro

6

s

10th

IS

20th

25th

30th

35

40

45

SO

SS

60

65

7

630096

phénol (melting point 114 ° C), 2,4-dinitrophenol (melting syl derivative is blocked. The degree of m substitution is point 113 ° C), 1-naphthol (melting point 94 ° C) and negligible.

2-Naphthol (melting point 122 ° C) makes the solvent. In the nitronium tetrafluoroborate process, the mixture is homogeneous and prevents charring. The preferred reaction time is 0.25 to 1 hour and the acetyl derivative is preferred. A suitable reaction temperature of about 25 ° C. is also suitable as solvent. The molar ratio of the above catalysts which is liquid in the reaction of nitronium tetrafluoroborate to acetylated glycoside, e.g. Titanium (IV) chloride (boiling between 1: 1 and 20: 1, with a ratio of 10: 1

point 136 ° C) or tin (IV) chloride (boiling point 114 ° C). is preferred to fully introduce a nitro

If desired, the product can ensure the reaction with the group. Besides the preferred dichloro-

Phenol reacetylated by the first stage method io methane can be used as chloroform and 1,2-dichloroethane to protect any free hydroxyl groups which solvents are used. The nitronium tetrafluoro-by side-deacetylation reactions during the introduction borate process may have arisen because of the ease with which the phenoxy group can be carried out. preferred.

The nitrided product obtained in the first process can be nitrated from the mixture of nitric acid, acetic acid and sulfur.

The direct use of 4-nitrophenol to produce ferrous acid can be isolated by the reaction mixture in 4-nitrophenylglycosides although water has been described (generally about 5 to 20 times its volume and can be used (TD Audichya, TR Ingle lot) poured and the crude product is either filtered off and JL Bose, Indian J. Chem. 9 (1971) 315, AP Jansen and or is extracted with chloroform (for nitronium tetra-PGAB Wydeveld, loc. cit.), but this is shown in Scheme II 20 fluoroborate process, the dichloromethane solution, which has been described in cold form and is saturated with oc- (4-nitrophenyl) tridecaacetylmaltote, is added over sodium

Traoside and a- (4-nitrophenyl) hexadecaacetyl maltopentaoside sulfate dried and evaporated, the crude glycoside leading process remains because of the easy implementation.

moves. The nitration can be carried out in a mixture of acetic acid and sulfuric acid with nitric acid or in dichloromethane for 2 seconds with a nitronium compound, e.g. Nitronium tetrafluoro-deacetylation reaction borate (N02 + BF4-) nitronium hexafluorophosphate (NCh + PFô ") The selective removal of O-acetyl groups from one or nitronium trifluoromethanesulfonate (NOa + CFìSOs) acetylated polyol derivative is preferably carried out using either a L catalytic converter Amount of sodium methoxide (in general Fieser and M. Fieser "Reagents for Organic Synthesis" 5 30 mean 0.01 to 0.1 molar equivalent) in methanol or (1975) 477, Wiley-Interscience, New York, described by a solution of anhydrous ammonia in methanol.

Nitronium tetrafluoroborate is preferred and can be used in addition to the preferred sodium methoxide

Description of this aspect of the method. In the first other lower alkali alkoxides, e.g. Potassium methoxide, a solution of the phenylacetylated glyco-sodium and potassium ethoxide and potassium t-butoxide, which are contained in the sids in a mixture of sulfuric acid and acetic acid at 35 corresponding alcohol, are used. 0 ° to 25 ° C with a 5- to 30-fold molar excess. These deacetylation reactions take place easily at temperatures of 70% nitric acid, which is dissolved in acetic acid, from 0 ° to 25 ° C within 12 to 24 hours,

treated. The nitric acid is preferably in a. The deacetylated product is by evaporating the alcohol.

Amount of 10 to 20 molar equivalents per molar hols isolated from inorganic ions by passage

Equivalent to the acetyl derivative used. The reaction temperature is freed by an acidic ion exchange column (if the temperature is between 0 ° and 25 ° C (preferably about 0 ° C), if desired) and from a suitable solvent, e.g. recrystallized by a further nitration of the aromatic ring and methanol or ethanol. Another one

Cleavage of the ester and glycoside bonds largely from deacetylation processes will switch on a 3% solution. The reaction time can be 1 to 10 hours. Hydrogen chloride in methanol (LF and M. Fieser “Reagents (preferably about 4 hours), but one should complete the reaction as completely as possible, 45 for Organic Syntheses”, page 11, Wiley, New York 1967) Let the above products run for a period of 4 to 24 without forming the above temperatures from 0 ° to 25 ° C. The nitriding hours are used. This is particularly advantageous for Dini's aromatic glycoside acetates, which are derived from the above in troverbindungen.

Connection with the displacement reaction mentioned In an alternative synthesis one of the phenols is used

Phenols have been prepared following the usual 50 displacement of halogen from tridecaacetylmaltotriosyl

o-p substitution scheme, whereby the p-position is favored or hexadecaacetylmaltopentaosyl chloride or bromide, unless it is used by another group as in 4-Kreder the following structure:

X = Cl or Br

630096

These halides are prepared either by treating the fully acetylated oligosaccharide with anhydrous hydrogen halide, solutions of hydrogen halide in mixtures of acetic anhydride and acetic acid, solutions of aluminum chloride and phosphorus pentachloride or of titanium tetrachloride in chloroform or from the oligosaccharide itself by treatment with acetyl chloride (W. Pigman, loc. cit., pages 150-151).

The halogen atom is either with phenol or a substituted phenol in the presence of a halogen acceptor such as silver (I) oxide (Ag20), silver (I) carbonate (Ag2C03), mercury (II) acetate (Hg (CH3COO) 2) or with Iron (III) chloride (Koenigs-Knorr reaction) or displaced with the sodium or potassium salt of phenol (W.Pigman, loc. Cit., Pages 194-198). The rest of the synthesis is the same as described above.

The course of all the reactions described above can be followed by thin layer chromatography (DSC) on silica gel in a suitable solvent system or by NMR spectroscopy. The purity of the Pro15

20th

Products of the reactions can be determined by high-performance liquid chromatography (HPLC), by plarimetry and by UV spectroscopy and high-frequency (220 MHz) NMR spectroscopy.

It has been found that the nitroaromatic glycosides according to the invention are advantageous substrates for the determination of a-amylase in serum. The determination method is illustrated in Scheme I for the preferred compounds according to the invention, a- (4-nitrophenyl) maltotetraoside and a- (4-nitrophenyl) maltopentaoside. The a-amylase in serum converts these two compounds into a mixture of G2 or G3 and a- (4-NytrophenyI) maltoside. The latter is then hydrolyzed to glucose and 4-nytrophenyl with a-maltase. Treatment with dilute alkali forms the 4-nytrophenolate anion, which can be identified spectroscopically and can be distinguished from any unreacted glycoside and can be related to the concentration of a-amylase in the serum.

Scheme I: Determination of a-amylase in serum

CKjOH

A, max 290-305 nm

R = 4-02NCeH4 n = 2 a- (4-nitrophenyl) maltotetraoside n = 3 a- (4-nitrophenyl) maltopentaoside la-amylase

CHoOH

G2 or g3 +

R = 4-O2NC6H4 n = O a- (4-nitrophenyl) maltoside a-maltase

V

4 or 5 Gì +

HO-V 7-N02

4-nitrophenol

OH-

0— (f y — N0j

4-nitrophenolate anion A, max410nm

9 630096

The nitroaromatic glycosides according to the invention amide adenine dinucleotide phosphate).

have the following advantages in determining the a-amy- Several embodiments of the invention are described in the serum: According to US Pat. No. 3,879,263, the a-amy- following examples are described. All quantities in the serum lasing content of the glucose parts and percentages formed from G4 or Gs relate to the weight, if related. Accordingly, the glucose in the s need not be specified otherwise. The chemical shifts in

Serum prior to the determination chromatographically from the nuclear magnetic resonance spectrum of the protons ('H

Sample to be removed. This requires the time required for NMR), unless otherwise stated, in parts per mil

the preparation of the sample and additional equipment. Lion's inner tetramethylsilane in chloroform-d (CDCb)

Use of the compounds according to the invention is indicated. The qualitative 1 H NMR results of the a-amylase content in the serum on the nitrophenols were determined at 60 MHz. More accurate measurements were made at 220

related, made regardless of the glucose content in the serum from MHz. The thin-layer chromatograms of the nitroaromatic glycosides of G4, Gs or G6 (DSC) were obtained on silica gel using 250 nm

have been put. This means that not only the chromato plates for the analysis work and 2 mm plates for the pre-

Graphics system for removing the serum glucose superfluous, parative work added. The high-performance liquid

but also the detection system by replacing the Hexaki 15 chromatography (HPLC) was combined with the device "Du Pont nose-ATP-NADP unit by diluted alkali 830" for the analysis work and with the device "Du Pont fold (ATP-NADP Adenosine triphosphate and nicotine 841 »is carried out for preparative work.

example 1

A. Preparation of maltotetraose-β-tetradecaacetate

CH20AC

k OAc A

AcO ^ j 1 O

OAc

CHo0Ac

0-OAc.

CH? OAc - Q CAc l \ p | ^

OAc

A mixture of 10.0 g (15.0 mmol) of maltotetraose with an analytically confirmed structure, 10.0 g (0.15 mol) of anhydrous sodium acetate and 50 ml of acetic anhydride was stirred at 100 ° C. for 2 hours and then poured into 300 ml of ice water . After 48 hours at 5 ° C, the colorless crystalline mass was filtered off and air-dried to give 20.96 g of crude product. This product was recrystallized from 40 ml of methanol, giving 18.18 g (14.49 mmol, 96%) of crystalline maltotetraose-β-tetradeca acetate in two crystals of 2.98 g and 15.20 g. The first crystals had a melting point of 124 to 126 ° C. Its structure was confirmed by the following values:

Vmax (CHCb) 1750, 1370, 1230 and 1030 cm-1; W (EtOH) 210nm (8 740); [a] g5 ° + 104 ° (c 1.03 CHCb); 'H NMR (220

MHz); 8 5.76 (d J = 7) (Hia), 5.43-5.25 (multiple groups of multiplets) 27H (OCH, OCH2) and 2.19-2.00 (series of singlets) 42H (COCH3) .

Elemental analysis for C52H70O35:

Calc .: C 49.76; H 5.62

Found: C 49.08; H 5.76

C 49.24; H 5.73

In several further experiments up to twice the above scale, the product yield after ss of recrystallization was 72-77%. The melting point was 122-128 ° C.

B. Preparation of the a- and ß-phenyltridecaacetylmaltotetraoside

CH ^ OAc

OAc

CBpOAc

OAc

OAc

OAc OAc

ZnCh

OH

630096

10th

CH20Ac f (oAc ^>! J '

o j— {o - J

OAc

CIloOAc

./"À,

"Where-

OAc

CH ^ OAc

A mixture of 11.0 g (8.77 mmol) of maltotetraose-β-tetradecaacetate, prepared according to section (A), 8.0 g (85 mmol) of phenol and 2.0 g (14 mmol) of anhydrous zinc chloride was gently heated, until it became thin, and then mechanically stirred at 100 ° C for 3 hours. The mixture was diluted with water and benzene and separated. The benzene layer was successively extracted three times with 50 ml of 5% sodium hydroxide and twice with 50 ml of saturated aqueous sodium chloride solution, dried and evaporated, 10.09 g (7.8 mmol, 89%) of the crude phenyl derivative in the form of a yellow crystalline Solid were obtained. This material was purified by preparative thin layer chromatography and high performance liquid chromatography as follows:

A total of 3.06 g of the crude phenyl derivative was placed on 14 2 mm plates for preparative thin layer chromatography and developed three times with a mixture of benzene and methanol (95: 5). The band at Rf 0.13-0.27 was extracted with chloroform and methanol to give 0.90 g (29%) of product which was recrystallized from ethanol with 0.58 g of a mixture of a- and ß- Phe-

10th

nyltridecaacetylmaltotetraosiden were obtained. Analytical HPLC (polar silicone microspheres) showed a product with a retention time of 8.99 minutes and a smaller impurity (2.4%) at 8.23 minutes. The structure of the crystalline phenyltridecaacetylmaltotetraoside was confirmed by the following values:

Vmax (CHCb) 1745, 1595, 1585, 1365, 1225 and 1030 cm- '; ? lmax (EtOH) 273 nm (s 800), 266 (960), 260 (730), 210 (7860); [a] g5 ° + 132 ° (c 1.00 CHCb); 'H NMR (220 MHz), Ô 7.37-7.25 (m) 2H, 7.14-6.95 (m) 3H (CeHs), 5.00-3.86 (m) 28H (OCH, OCH2) and 2.19-1.97 ppm (set of singlets), 39H (COCHa).

Elemental analysis for C56H72Q34:

15

Calc .: C 52.17; Found: C 51.44; H 5.35;

H 5.63 H 51.94; H 5.60;

H 51.60 H 5.39

25th

30th

A total of 4.25 g of the crude phenyl derivative was also purified by preparative HPLC on a Spherosil (44-50) column of 1 mx 23 mm, using a 1: 1 mixture of pentane and dioxane (water content 1.5% ) was eluted and 1.07 g (yield 25%) of colorless crystalline phenyltridecaacetylmaltotetraoside melting point 82-83 ° C were obtained. According to the spectral and chromatographic data, this product was identical to the material purified by thin-layer chromatography. Analysis by HPLC showed that this sample was 99.7% pure.

C. Preparation of a- and ß-Phenyltridecaacetylmaltotetraosiden (Alternative Procedure)

AcO

CHjjOAc oi — r 0

OAc

OAc

OAc

CII? 0Ac

4T

Oll

ZnCh / Ac0H / Ac20

OAc

CHpOAc

/ \

K OAc A

AcO 1 1 0

OAc

CHpOAc

Np ^ 1-0.

OAc

CHpOAc

OAc V '

2.75 g (2.19 mmol) of maltotetraose-β-tetradecaacetate and 2.27 g (24.2 mmol) of phenol were mixed in a three-necked flask under nitrogen. A solution of 0.55 g of zinc chloride in 2.0 ml of a mixture of acetic acid and acetic anhydride (95: 5) was added and the reaction mixture was slowly warmed. When the mixture became homogeneous (temperature about 45 ° C), the internal pressure was gradually lowered to 23 mm Hg and the reaction mixture was stirred at 100 ° C for 2.5 hours. The mixture was transferred to a separatory funnel using 200 ml of warm benzene. The cooled mixture was washed twice with 18% aqueous sodium chloride, twice with 50 ml 2.5% sodium hydroxide and twice with 30 ml saturated sodium chloride solution. The organic layer was dried over sodium sulfate and evaporated and the residue treated with 3.0 g of anhydrous sodium acetate and 15 ml of acetic anhydride. The resulting mixture was heated to 120 ° C for 1.0 hour, cooled and treated with 200 ml of ice water. Any unprotected hydroxyl groups that arise during treatment with phenol were acetylated again. The solid that formed on standing at 0 ° C was filtered off and dried. Yield 55 2.85 g. This material was chromatographed on silica gel (Mallinckrodt SilicAR CC-7). Elution with a mixture of benzene and methanol (97: 3) gave a total of 2.07 g (71%) of pure phenyltridecaacetylmaltotetraoside after recrystallization from ethanol. This material 60 had a melting point of 112-117 ° C. Its structure has been confirmed by the following data:

[a] jf + 135 ° (c 1.0 CHCb),> H NMR (220 MHz) 5 7.38-6.96 (m) 5H (C6H5), 5.77 (dd J = 9.5 Hz) (Hia) and 5.61 (d J = 4 es Hz) (Hiß) 1H, 5.45-3.82 (series of multiplets) 27H (OCH, OCH2) and 2.24-1.95 ppm (series of Singletts) 39H (COCH3); the integration was consistent with a 75:25 mixture of a: β-phenyltridecaacetylmaltotetraosiden.

11 630 096

D. Preparation of a- and β- (4-Nytrophenyl) tridecaacetylmaltotetraoside (Nitric Acid Process)

CILOAc aÌWo-

OAc

CHpOAc

^ - <1

nÇAc /

f— | 0

OAc i

Cll ,, OAc

J ^ -Co

OAc

HN03, H2SO4

■

HOAc

CII? OAc

'Ö II

0 ^ - 0 - J

CHpOAc

AcO

A mixture of 2.0 ml of acetic acid, 1.0 ml of acetic anhydride and 0.38 g (0.29 mmol) of a mixture of a- and ° C cooled and treated with a mixture of 1.0 g of sulfuric acid and 2.0 g of acetic acid and then with a mixture of 0.5 g of 70% nitric acid and 1.0 ml of acetic acid. The mixture was stirred at 25 ° C for 4 hours, poured into 25 ml of ice water and filtered to give 0.40 g of an almost colorless solid. This sample consisted of approximately 50% of the 4-nitro-phenyl derivative, as shown by the value of ^ max (EtOH) of 290 nm (£ 3100).

The reaction was repeated on a larger scale (5 g of crude phenyltridecaacetyl maltotetraoside) to give 4.35 g (84%) of the crude nitro compound which was chromatographed on 200 g of silica gel (Mallinckrodt SilicAR CC-7). The product eluted with 4% methanol in benzene (2.27 g recovered) was also about 50% pure.

A 2.6 g sample of the crude precipitated nitro compound was prepared by preparative HPLC separation with

20 using a 2 m x 2.3 cm column of the product “Spherosil (44-50)”. The column was eluted with a 1: 1 mixture of pentane and dioxane (water content 1.5%). The fractions were examined by thin-layer chromatography and UV spectroscopy. 0.2557 g (0.19 mol, 8%) of pure α- and β- (nitrophenyl) tridecaacetylmaltotetraoside were obtained from the medium-sized sections. After recrystallization from 5 ml of ethanol, this material had a melting point of 112-114 ° C. The structure of the material was confirmed by the following values:

30th

Vmax (CHCb) 1745.1580, 1520, 1420, 1370 and 1220cm-1; A, max (EtOH) 290 nm (e 7180); [a], f + 156 ° (c 0.525 CHCb); 1H NMR (220 MHz), 8 8.26 (d J = 8) and 7.26 (d J = 8) 4H (CóHt), 5.75 ( m) IH (Hia and Hiß), 5.52-3.95 (series of Mul-35 tipletts) 27H (OCH, OCH2) and 2.25-2.01 ppm (series of singletts) 39H (COCH3).

Elemental analysis for C56H71NO36:

40 calc .: C 50.41; H 5.36 Found: C 49.95; H 5.54

E. a- and ß- (4-nitrophenyl) tridecaacetylmaltotetraoside (nitronium tetrafluoroborate process)

CHpCAc

CHpOAc

\ fVo Jxî-ro <Q>

OAc OAc

OAc

CHpOAc

ICOAc

AcO i

A slurry of 1.91 g (14.5 mmol) of nitronium tetrafluoroborate in 8 ml of dry methylene chloride was treated with a solution of 1.24 g (0.96 mmol) of phenyltridecaacetylmaltotetraoside from part (C) in 10 ml of dichloromethane. The resulting mixture was stirred under an argon atmosphere for 20 minutes. The mixture was added to 60 ml of ice water, separated and washed with cold aqueous saturated sodium chloride solution. The organic part

0-

CllpOAc &

OAc

N02 + BF4-CH2CI2

CH ~ 0fcc

Kpvi

OAc

VQ,

J?

was dried over magnesium sulfate and evaporated under reduced pressure to give 1.30 g of a- and β- (4-nitro-phenyl) tridecaacetylmaltotetraoside in the form of a yellow glassy solid. Half of this product was recrystallized from ethanol to give 0.48 g (75%) of an off-white solid of melting point 116-119 ° C. The structure of the product was confirmed by the following data:

630096

12th

'H NMR (220 MHz), ô 7.75 (AA'BB') 4 H (CeH4), 5.75 (d J = (COCHs); W (EtOH) 288 nm (e 5840); [a] g ° + 152 ° (c 0.5 4 Hz) (hot) and 5.81-3.86 (series of multiplets) 28H (Hia, CHCb).

OCH, OCH2) and 2.25-1.95 (set of singlets) 39H

F. Preparation of a- and ß- (4-nitrophenyl) maltotetraoside

CHgOAc

OAc OAc t / * ~ q

CHpOAc

CHpOAc for \ A rv '

NaOCHs / CHsOH

NO.

CHgOll

- "fò-P

OH

CH-OH

OH

CH-OH

£> o

OH

tfo.

-J 2nd

A mixture of 6.76 g (5.1 mmol) of (4-nitrophenyl) tride-caacetylmaltotetraoside from part (D), which had been subjected to column chromatography, 50 ml of methanol and 50 mg of sodium methoxide was kept at 25 ° C. overnight . The mixture was filtered to remove a small amount of a yellow solid. The filtrate was evaporated to leave 3.17 g (4.02 mmol, 79%) of a yellow solid. 3 g of this sample was purified by preparative thin layer chromatography on 11 plates by development in a mixture of chloroform, acetic acid and water (3: 3.5: 0.5). 0.5027 g material from the gang

Rf 0.16-0.29 consisted of a mixture of a- and ß- (4-

2s nitrophenyl) maltotetraoside with a purity of about 50% (according to the UV spectrum). The product was a yellow solid, melting point 68-70 ° C. Its structure was confirmed by the following values:

Vmax (Nujol) 3300 and 1020 cm- '; W (H2O) 305 nm (e 3740);

30 [<x & 5 ° + 104 ° (c 1.06 H2O); 'H NMR (220 MHz), S 8.25 (d J = 10) and 7.35 (d J = 10) (CeHt), 7.91 (t J = 7) and 7.70 (t J = 7) (other aromatic material), 5.88-5.79 (m) (Hia and Hiß), 5.30 (m) (OCHO), 4.98 (s) (HOD) and 4.02-3, 48 ppm (m) (OCH, OCH2).

G. (4-nitrophenyl) maltotetraoside (alternative method) CII ^ OAc CHpOAc CHpOAc

OAc OAc OAc

NaOCHs / CHsOH

CH..0H

"O <lVo

OH

A mixture of 250 mg (0.19 mmol) (4-nitrophenyl) tri-decaacetylmaltotetraoside from part (E) in 2.5 ml of methanol was mixed with 3.3 ml of a 7.8 x 10 "3-molar solution of sodium methoxide in Treated methanol and stirred at room temperature for 17 hours. The solvent was removed under reduced pressure. The residue dissolved in the minimum amount of methanol (2 ml) was added dropwise to 40 ml of ether. The resulting yellow powder was separated, dissolved in methanol and dissolved by a Column of the ion exchange resin "Sephadex LH-20" of 2.5 x 20 cm passed, using methanol as the eluent, giving 134 mg (90%) of solid product which was chromatographed again, with 90 mg (60%) of a medium Were obtained, the structure of which by the

^ 2

55 the following values were confirmed:

Uax (H2O) 303 nm (e 6350), 217 (e 5000); > H NMR (220 MHz), 8 (CD3OD) 7.77 (AA 'BB', Jab = 9.5 Hz) 4H (CsH »), 5.69 (d J = 4 Hz) 1H (hot), 5 , 27 = 5.09 (m) 3H (OCHO), 4.87 (s) 13H (HOD), 4.20-4.05 (m) (minor contamination) 60 and 4.01-3.40 ppm ( m) (OCH, OCH2).

H. (4-nitrophenyl) maltotetraoside (alternative method) In the manner described in part (G), a crude sample of (4-nitrophenyl) maltotetraoside was prepared with the following 65 key figures:? Unax (H2O) 302 nm (e 5900), 217 (4650); [ccß5 ° + 158 ° (c 1.1, CH3OH). Part of this sample was purified by fractionation on a jx Bondapak / carbohydrate column (Waters Associates) using a

13

630096

Mixture of acetonitrile and water (87:13) was eluted. The purified main component of the sample obtained after freeze drying showed Xmax (H2O) 302 nm (e 9425), 220 (6500).

Example 2

A. Preparation of maltopentaose-β-heptadecaacetate

CHo0H

CH-Oll

011

CH, 0H

Na0Ac / Ac20

AcO

A mixture of 2.00 g (2.41 mmol) of maltopentaose, 2.0 g (30 mmol) of sodium acetate and 20 ml of acetic anhydride was heated to 130 ° C for 2.0 hours. The cooled mixture was added to 75 g of ice and allowed to stand at 0 ° C. The solid formed was pulverized and filtered to give 3.35 g (90%) of an off-white solid which was recrystallized from 40 ml of ethanol. 3.16 g (85%) of maltopentaose-.beta.-heptadeca acetate of melting point 117 to 120 ° C. were obtained. The structure was confirmed by the following key figures:

[a] lf + 122 ° (c 0.7, CHCb); 'H NMR (60 MHz) 5 5.85-3.78 (m) 35H (OCH, OCH2), and 2.34-1.83 ppm (series of 25 singlets) 51H (COCH3); 'H NMR 220 MHz) 5 5.76 ppm (d J = 8 Hz), 1H (Hia);

Elemental analysis for C64H86O43:

30 calc .: C 49.80; H 5.62 Found: C 49.11; H 5.54 C 49.28; H 5.71

B. Preparation of a- and β-phenylhexadecaacetyl maltopentaoside

CHpOAc

CII ,, 0Ac

CHpOAc

* 0Ac OAc OAc

OH

ZnCk

>

AcO

CHo0Ac OAc

CHpOAc

^ ° <Q>

OAc N '

A mixture of 3.00 g (1.95 mmol) of maltopentaose-β-heptadecaacetate from part (A), 1.95 g (20.8 mmol) of phenol and 0.46 g (3.38 mmol) of anhydrous zinc chloride became 3 Heated at 100 ° C under an argon atmosphere. The cooled mixture was taken up in 200 ml of dichloromethane and washed twice with 30 ml of water, twice with 25 ml of 5% sodium hydroxide and twice with 25 ml of saline. The organic layer was dried over magnesium sulfate and concentrated under reduced pressure to give a yellow powder which was treated with 3.0 g of sodium acetate and 15 ml of acetic anhydride and then heated at 125 ° C for 2.0 hours to remove any free hydroxyl groups to acetylate. The cooled mixture was added to 125 g of ice water and allowed to stand at 5 ° C. The solid a- and ß-phenylhexadecaacetylmaltopentaoside formed in this process was pulverized, separated and air-dried, 2.45 g of a white 55

60

65

powder was obtained. This material was chromatographed on 150 g of silica gel, eluting with a mixture of benzene and methanol (97: 3). The 'H NMR spectrum (220 MHz) of the residue from a medium section showed 8 7.38 to 7.27 (m) 2H and 7.17-6.97 (m) 3H (CsHs), 5.77 ( dd J = 9.5 Hz), 5.63 (d J = 4 Hz) (hip), 5.82-3.83 (series of multiplets) 35H (OCH, OCH2) and 2.23-1.93 ppm (Set of singlets) 48H (CH3CO).

Pure phenylhexadecaacetylmaltopentaoside was obtained from the above solid product by HPLC using> 40 | i silica gel column columns. obtained with a mobile phase from 1% methanol, 49% pentane and 50% dichloromethane. Integration of the 220 MHz 'H NMR spectrum showed that this material consisted of approximately 75% a-phenylhexadecaacetylmaltopentaoside and 25% ß-phenylhexadecaacetylmaltopentaoside.

630096

14

C. Preparation of a- and ß-phenylhexadecaacetyl maltopentaoside (alternative method) CHo <

CU OAc

CHpOAc

/ l - J— ° X 0AC OAc r \

OH

ZnCh / Ac0H / Ac20

OAc

CIUOAc

OAc OAc

CHgCAc

A mixture of 3.00 g (1.95 mmol) of maltopentaose-β-heptadecaacetate from part (A) and 1.95 g (20.8 mmol) of phenol in a three-necked flask under nitrogen was mixed with a solution of 0.49 g of zinc chloride treated in 2.0 ml of a 95: 5 mixture of acetic acid and acetic anhydride. The reaction mixture was slowly warmed. When the reaction mixture became homogeneous, the internal pressure was gradually lowered to 23 mm Hg and the mixture was stirred at 100 ° C for 2.5 hours. The residue was treated with 200 ml of benzene and 18% aqueous sodium chloride solution. The organic phase was washed twice with 50 ml of 2.5% sodium hydroxide solution and twice with 50 ml of saturated sodium chloride solution. The organic layer was dried over sodium sulfate and evaporated. The residue was treated with 3.0 g of anhydrous sodium acetate and 15 ml of acetic anhydride. The mixture became 1.0

Heated to 120 ° C for one hour, cooled and treated with 200 ml of ice water 20. The solid product which formed on standing at 0 ° C was pulverized, filtered off and air dried to obtain 2.97 g of an off-white powder. This material was chromatographed on a silica gel column, eluting with a 97: 3 mixture of benzene 25 and methanol. A total of 2.00 g (65%) of pure a- and β-phenylhexadecaacetylmaltopentaoside were obtained with a melting point of 119-125 ° C. [a] β ° + 137 ° (c 1.0, chloroform).

30 Elemental Analysis for CósHssCte:

Calc .: C 51.77; H 5.62 Found: C 51.62; H 5.70 C 51.13; H 5.47

D. Preparation of (4-nitrophenyl) hexadecaacetyl maltopentaoside

OlgOAc Clf ^ OAc OAc OAc

CH-OAc

./•""A

r AC. .

OAc

CHpOAc

+ NO2 + BF4-

OAc

CHpOAc

CH ^ OAc

AC „kfVo-Mi-i ^ F-

CH2CI2

A slurry of 0.85 g (6.4 mmol) of nitronium te was obtained. The structure of the product was confirmed by the trafluoroborate in 10 ml dry dichloromethane under the following values:

an argon atmosphere with a solution of 1.00 g (0.64

mmol) phenylhexadecaacetylmaltopentaoside from part (C) in "> ^ £ ax0H 290 nm (e 5550), 210 nm (e 7750); 'H-NMR spectrum treated 12 ml dichloromethane. The resulting mixture (220 MHz) 8 7.75 (AA'BB ', Jab = 9.5 Hz) 4H (CöHj), 5.75 (d was stirred for 20 minutes at room temperature and under J = 4 Hz) (hot) , 5.78-3.85 (series of multiplets) (OCH, stirring in 70 ml of cold saturated sodium chloride solution OCH2) and 2.22-1.93 ppm (series of singlets) 48H

poured. The deposited organic layer was over (CH3CO).

Magnesium sulfate dried. By evaporating the ß by two recrystallizations of the crude (4-nitro-

Solvent under reduced pressure gave 1.05 g of phenyl) hexadecaacetyl maltopentaoside from ethanol. A yellow solid was obtained, which from 30 ml of ethanol gave a sample with A, max (CH3OH) 290 nm (e 6650) and 212 (7700)

was recrystallized to obtain 0.87 g of an off-white powder.

15

E. Preparation of (4-nitrophenyl) maltopentaoside

630 096

CJ ^ OAc CHoOûc CHpOAc

J- \ J- \ _ 1

OAc OAc OAc

NaOCHs / CHsOH

CH? 0H

■ x0H

HO} —H 0 • OH

CllpCH

| 0 ..

OH

CH? CH

/ "X,

N6,

OH

A solution of 430 mg (0.27 mmol) (4-nitro-phenyl) hexadecaacetyl maltopentaoside from part (D) in 5 ml of methanol was treated with 4.7 ml of a 7.8 x 10-4 molar solution of sodium methoxide in methanol . The mixture was stirred at room temperature for 17.5 hours. The solution was concentrated under flowing dry nitrogen. The residue was treated with 40 ml of ether with stirring to precipitate a solid, which was filtered off and air dried. This gave 247 mg (98%) of a slightly yellow powder, /.max (H2O) 303 nm (e 5980), 215 (5530). HPLC (| i Bondapak / coal hydrate from Waters Associates, 80% acetonitrile / water, 254 nm, UV detector) of the product showed two peaks of the same intensity with retention times of 10.9 and 12.3 minutes corresponding to 4- ( Nitrophenyl) maltopentaoside and an unidentified material.

Examples 3 to 32 The experiments described in Examples 1 (B), 1 (C), 2 (B) and 2 (C) were repeated, but instead of the phenol used in the experiments described in Examples 1 and 2 the substituted phenols mentioned below in column A of Table II were used as reactants. The a- (substituted aryl) polyacetylglycosides mentioned in column B were obtained. By nitrating these products in the manner described in Examples 1 (D), 1 (E) and 2 (D), the a- (substituted nitroaryl) polyacetylglycosides mentioned in column C became

20 received. Deacetylation of the products mentioned in column C in the manner described in Examples 1 (F), 1 (G), 1 (H) and 2 (E) gave the a- (substituted nitroaryl) glycosides mentioned in column D. .

Maltohexaose-ß-eicosaacetat (GôAc2o) can be prepared by acety-lation of maltohexaose in the manner described in Examples 1 (A) and 2 (A). Maltotetraose, maltopentaose and maltohexaose are produced in the manner already described.

From the starting materials listed in column A of Table II, the following compounds are commercially available: 2-, 3- and 4-methylphenol (o-, m- and p-cresol), 2-, 3- and 4- Chlorophenol, 2-fluorophenol, 3-bromophenol, 4-iodophenol, 2-, 3- and 4-nitrophenol, 4-methoxyphenol, 2-isopropyl-5-methylphenol (thymol), methyl salicylate, 3s 3-ethylphenol, 4-t- Butylphenol, 1- and 2-naphthol, 4-chloro-1-naphthol and 4-hydroxybenzoic acid (for the production of hexyl 4-hydroxybenzoate). The following starting materials are all described in the «Dictionary of Organic Compounds», 4th edition, edited by I. Heilborn, Oxford 40 University Press (1965) on the pages mentioned: 2-methyl-5-isopropylphenol (Carvacrol, page 568), 2-methoxyphenol (Guaiacol, page 1549) and 3-methoxyphenol (page 2857). Hexyl 4-hydroxybenzoate can be prepared by acid catalyzed esterification of 4-hydroxybenzoic acid 45 with 1-hexanol following the procedure for methyl salicylate by A.I. Vogel “Practical Organic Chemistry”, 3rd edition, Longmans Green, London 1959, page 782.

Table II

example

Column A

Column B

3rd

2-CH3C6H4OH

G4 (Ac) l3 (2-CH3C6H4)

4th

3-CH3C6H40H

G4 (Ac) i3 (3-CH3C «H4)

5

4-CH3C6H4OH

G5 (Ac) i6 (4-CH3CeH4)

6

2-C1C6H40H

G5 (Ac) i6 (2-ClC6H4)

7

3-C1C6H40H

Gs (Ac) i6 (3-ClC6H4)

8th

4-C1C6H40H

G4 (AC) 13 (4-C1C6H4)

9

2-FC6H40H

G4 (Ac) i3 (2-FC6Ht)

10th

3-BrC6H40H

Gs (Ac) i6 (3-BrC6H4)

11

4-IC6H40H

G4 (AC) 13 (4-IC6H4)

12

2-O2NC6H4OH

G4 (Ac) l3 (2-02NC6H4)

13

3-O2NC6H4OH

Gs (Ac) i6 (3-02NC6H4)

14

4-O2NC6H4OH

Gs (Ac) l6 (4-02NC6H4)

15

2-CH3OC6H4OH

G4 (Ac) i3 (2-CH30CóH4)

16

3-CH3OC6H4OH

Gs (Ac) i6 (3-CH30C6H4)

17th

4-CH3OC6H4OH

G4 (AC) 13 (4-CH30C6H4)

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Table II (continued)

630096

example

Column C

Column D

28

G6 (Ac) i9 (2-C1-4-02NC6H3)

G6 (2-Cl-4-02NC6H3)

29

G6 (Ac) 19 (3,5- (02N) 2C6H3)

G6 (3.5- (02N) 2C6H3)

30th

G6 (AC) 19 (2-CH30-4 ~ 02NC6H3)

G6 (2-CH20-4-02NC6H3)

31

G6 (Ac) i9 (4-02N-1 -C ioH?)

G6 (4-02N-1-CioH7)

32

G6 (Ac) i9 (4-02NCeH4)

G6 (4-02NCeH4)

Claims (10)

630096 2 PATENT CLAIMS 1. Process for the preparation of nitroaromatic glycosides of the formula (I) ch2oh CHjOH ch2oh (I), wherein n is an integer of 2, 3 or 4 and R is a substituted aromatic radical of the formula X - or is, wherein X and Y are independently H, no2, halogen, alkyl having 1 to 4 carbon atoms, OR1 or co2r1 Provided that at least one of the radicals X and Y stands for no2, characterized in that one with R1 is an alkyl radical with 1 to 6 carbon atoms with the a) acetylated glycosides of the formula CH-OA r 2 C AcO QAc CH-OA ■ 2 c \ OAc OAc n CH-OÄ 2 C K wherein Ac is an acetyl radical and n has the meaning given above, with a phenol of the formula y or or at most one of the substituents X and Y is no2, in the presence of a catalyst at a temperature in the range from 80 to 120 ° C., b) nitriding the product formed in step a) by using it I) nitric acid contained in a mixture of acetic acid and sulfuric acid, or II) a nitronium compound selected from nitronium tetrafluoroborate, nitronium hexafluorophosphate and nitronium trifluoromethanesulfonate and contained in dichloromethane, chloroform or 1,2-di-chloroethane and c) the product obtained in stage b) is deacetylated by ss it with I) a catalytic amount of an alkali metal lower alkoxide contained in the corresponding alcohol or 60 II) a solution of anhydrous ammonia or HCl in methanol. 2. The method according to claim 1, characterized in that p-toluenesulfonic acid as the catalyst in stage a) 65 or an anhydrous covalent metal chloride, in particular zinc chloride, and the reaction in stage a) is carried out at a temperature in the range from 100 ° to 110 ° C. 3. The method according to any one of claims 1 or 2, characterized 3rd 630 096 characterized in that the nitration reaction in step b) is carried out by combining the product of step a) with nitronium tetrafluoroborate, which is contained in dichloromethane, chloroform or 1,2-dichloroethane, at a temperature of about 25 ° C., the Molar ratio of nitronium tetrafluoroborate to the product of stage a) is in the range from 1: 1 to 20: 1, in particular 10: 1. 4. The method according to any one of claims 1 to 3, characterized in that one carries out the deacetylation reaction in step c) by the product of step b) with 0.01 to 0.1 molar equivalents of sodium methoxide, which is contained in methanol , at a temperature in the range of about 0 ° to 25 ° C. 5. The method according to claim 1, characterized in that R in the product compounds of formula (I) contained therein represents. 6. The method according to any one of claims 1 or 5, characterized in that X in the resulting product compounds of formula (I) is no2 and Y is H. 7. The method according to claim 1, characterized in that R in the resulting product compounds of formula (I) is a 4-nitrophenyl radical. 8. Compounds of formula (I), prepared by the process according to claim 1. 9. Compounds according to claim 8, manufactured ch2oh 15 according to the method according to patent claims 5 to 7. 10. Use of the compounds of formula (I) according to claim 8 for the determination of a-amylase in biological media. 20th