Las nuevas fuentes del conocimiento

o-Gluoosone was prepared in poor yield by the decomposi­ tion of D-glucose bishydrazone with benzaldehyde; severe looses were incurred during the purification of the crude product.

Brull (1936) described the decomposition of D-glucose phenylosazone with an excess of pyruvic acid in hot aqueous sol­ ution; pyruvic acid phenylhydrazone separated on cooling ^nd was filtered off ^vhile excess pyruvic acid was removed by extrac­

tion with ether. Evaporation of the aqueous solution, after decolorisation with charcoal, gave D-glucosone in 40% yield. The present author has obtained D-glucosone, 3-’i)’*a:cthyl u-gluo- osone, D-galactosone, and L-gulosone in 60-65% yields by a mod­ ification of the above method. The corresponding phenylosazones, suspended in water, were decomposed with a little less than the calculated amount of pyruvic acid by heating at 100° for 1-2

hours, and isolated as described above; Brull (1936) recommended a reaction time of half an hour. By use of an excess of phenyl­ osazone contamination of the osone products \ith unchanged

pyruvic acid, whioh is very soluble in water as well as in ether, was reduced to a minimum.

By the action of glyoxal on D-gluoose phenylosazone, following the procedure described for decomposition with pyruvic acid, crude D-gluooeone was obtained in 70% _{yield by}

the present author. However, it was demonstrated that the product was invariably contaminated with unchanged glyoxal, in spite of employing a slight excess of phenylosazone and allowing a reaction time of 2 hours; glyoxal was not removed by extrac­ tion with ether. In consequence, such a method is of no value for the preparation of osones for structural and metabolic

investigations without additional chromatographic purification or preparation of crystalline derivatives from whioh the osones may be regenerated (see Part II, 1.$.).

I

1

91

1.3# By Direct Oxidation of the Corresponding Aldose or Ketose. 1.3.1. Aotion of Cuprio Acetate.

o-bVuctose was oxidised with a saturated aqueous solution of oupric acetate at 50° for 30 hours according to the directions of Evans, Nicoll, Btrause & Waring (1928). After filtration from the precipitated cuprous oxide removal of inorganic contam­ inants was effected by treatment with hydrogen sulphide followed by passage of the solution through columns of ion-exchange

resins. The resulting solution gave qualitative tests for both osone and unchanged fructose; in consequence, the stiff syrup obtained by evaporation under reduced pressure was treated with dry acetone containing concentrated sulphuric acid (4%« by

volume); the syrupy products were isolated in the usual manner and extracted with hot water. From the syrupy residue tri-^- -isopropylidene o-gluoosone hydrate was obtained by crystallisa­ tion from methanol (see Iart II, 3.2.2.); from the aqueous extract 2%3-4:5~di-0-isopropylidene d-fructose separated on cooling. From l8g. of o-fructose 0.-35g. of the isopropylidene o-gluoosone derivative and 8.5g. of di-O-isopropylidene o-fruc­ tose were obtained. Calculated on the basis of a 15% conversion of o-gluoosone to tri-Q-iaopropylidene o-glucosone hydrate these figures represent the formation of the osone in approximately 10% yield by the oxidation of d-fructose. Evans ^ al. (1928)

reported a 24% yield of osone, estimated as o-gluoose phenyl­ osazone. This confirmation of the low yield of osone by this method makes it obvious that the procedure has little preparative value.

o-Gluoose, L-Gorbose, and o-xylose were oxidised with cupric acetate according to the method of Weidenhagen (1937). Since the products of oxidation were shown to contain unchanged starting material as well as osones they were condensed with acetone and from the products di-Q-isopropylidene derivatives of of the un-oxidised sugars were removed by hot aqueous extraction. From the water-insoluble residues crystalline isopropylidene

derivatives of the corresponding osones, when o-gluoose and L-sorbose were oxidised, were obtained by crystallisation from methanol in yields representing a 40% conversion of the sugars

to the corresponding osones. The method of preparation may be carried out on a large scale and has provided a valuable source of the crystalline isopropylidene derivatives from which

the pure osones may be obtained by hydrolysis.

1.3.2. Aotion of Selenious Acid.

From o-fructose, by the method described by Dixon & Harrison (1932), o-gluoosone was obtained in 8% yield; Dixon & Harrison (1932) did not quote a yield. These workers claimed that o-gluGosone prepared in this manner regenerated the colour of Soniff* 8 reagent and formed an addition compound with sodium bisulphite; the present author has been unable to confirm these observations with gluoosone prepared either by this method or by any other.

95.

1.4. ladireot Syntheses of D-Qluoo#o^#.

1.4*1. Oxidation of 2;3%4:6-Tetra-0^Aoetyl-2~0xy-D-Qluoal.

2:5:426~T*tra-0-ao$tyl-2-oxy-D-gluoal was prepared from

2:5:426-tetra-0-ao@tyl-D-gluooayl bromide, in turn prepared

direotly from D-glucose by the method of Martos & Korosy (1950), by treatment with diethylamine according to the directions of Naurer (I929)* Oxidation of the u-glucal derivative with perbenzoic acid (Stacey & Turton, 1946) gave crystalline

2:3:4*6-tetra-0-acetyl D-glucosone hydrate (VII) in comparatively poor yield, a large proportion of tae starting material being recovered. The same o-glucosone derivative was also prepared by treatment of the non-crystalline products of the chlorination of tetra-O-aoetyl-2-oxy-D-glucal, in ethereal solution, with silver carbonate and a little water, as described by Maurer

(1929)* Stacey 6 Turton (1946) reported the product of per­ benzoic acid oxidation of the o-glucal derivative to differ from

that of Maurer (1929) in that it did not exhibit mutarotation and also with regard to melting point. however, by either method of preparation, the samples of 2:524:6-tetra-0^-acetyl

D-gluoosone hydrate obtained by the present author showed p.iys- ical properties identical with those recorded by Maurer (1929)

for this derivative. That the derivative contains an incipi- ently ionic hydrogen atom, as was claimed by Stacey & Turton (1946), was confirmed by titration with dilute alkali.

1 ^ c(oh)

H A

'(VII) (XXI)

):4:6-Tri-0-acetyl D-gluoosone hydrate (XII) was prep­ ared from the syrupy chlorination products of 2:3:4z6-tetra-0- -aoetyl-2-oxy-D-glucal by treatment with sodium bicarbonate and a small amount of water according to the method of Maurer

& Petsoh (19)1); the product showed physical properties iden­ tical with those given by these latter workers.

The present author has demonstrated unequivocally that

2:):4î6-tatra-0-aoetyl o-^luoosone hydrate and 5?4:6-tri-^- -acetyl o-gluoosone hydrate are indeed derivatives of o-gluco- sones such proof was not given by either Maurer and his

coworkers or Stacey & Turton (1946) (see Part I, 1.4.1.).

After short treatment with dilute alkali both derivatives red­ uced Tehling's solution at room temperature, readily formed

D-gluoose phenylosazone with phenylhydrazine in acetic acid solution, and gave a blue colour with Benedict's areenophospho- tungstic acid reagent in the presence of alkali-cyanide;

catalytic deaoetylation with metallic sodium in anhydrous methanol gave o-gluoosone, identified chromatographically on paper employing the upper layer of a ^-butanol-acetic acid- -water (4:1:$) mixture as developer and triphenyltétrazolium chloride as identification reagent; treatment of either deri­ vative with anhydrous acetone containing concentrated sulphuric acid gave a crystalline product identified as tri-Q-isopropyl- idene o-glucosone hydrate, a derivative also obtailed by

similar treatment of o-gluoosone (see Part II, ).2.2.)#

The preparation of u-gluoosone by deaoetylation of the crystalline acetates described above, although giving a product possessing a high degree of purity, involves the initial prep­ aration of four intermediates (tetra-O-acetyl-u-glucosyl

bromide, tetra-O-acetyl-2-oxy-D-glucal, the chlorination prod­ uct of the latter, and the desired u-gluoosone acetate), of which only the first may be obtained in good yield; in conse­ quence, alternative methods of preparation, such as decompo­ sition of the corresponding phenylosazone with benzaldeiiyde, are to be preferred.

95.

1.4.2. Oxidation of 2:)-4:5-Di~0-i8O^ropylidene u-fruotose.

Oxidation of the free primary alcohol group of 2:3-4î5- -dl-O-lGopropylldnne o-fruotoee (XCVI) to an aldehyde group should provide anjlnterestlng dl-O-lsopropylldene derivative of D-gluoo8one In the non-hydrated form possessing a free aldehyde group and a 2%6-fruotopyranose ring structure. however,

attempts by the present author to carry out such an oxidation and Isolate the product have met with no success.

Ô

(XCVI)

Thé use of selenlous acid for the contemplated oxidation Is precluded by the strong acidic conditions of such a method which would bring about hydrolysis of the Isopropylldene groups of both the substrate and the product. It has been shown that no oxidation of dl-O-lsopropylldmne u-fructose occurred on

treatment with cuprlc acetate in methanol; this observation indicates the necessity of the presence of a free hydroxyl group (or, perhaps, a free carbonyl group) adjacent to the hydroxyl group to be oxidised by this reagent. Such a structural envir­ onment is also considered to be required for the oxidation of hydroxyl groups by Fenton's reagent (see iart I, 1.3*1.)* and in consequence the use of this latter oxidant was not attempted.

From the attempted oxidation of the fructose derivative, in chloroform solution, with manganese dioxide at room tempera­ ture for 6 days the starting material was recovered almost quantitatively; the small, non-crystalline residue did not regenerate the colour of Schiff*s reagent, reduce Benedict's reagent, or form an addition compound with dimedone.

Oppenauer & Oberrauoh (1949) described the use of tert-butyl chromate, a strongly acidic reagent, in non-polar

organic solvents for the selective and almost quantitative oxidation of primary alcohol groups to aldehyde groups.

Attempts by the present author to oxidise di-Q-iaopropylidene D-fructose, in petroleum solution and using solid calcium carbonate as a buffer, with this reagent have met with little success; qualitative evidence of oxidation having occurred was obtained but isolation of the product was not achieved.

1.4.3. Oxidation of N-p-Tolyl-p-isoGlucosamine.

Weygand & Schaefer (1931) claimed that oxidation of N-j^-tolyl-o-mannamine (ji-j^-tolyl-l-amino-l-deeoxy o-mannitol)

(XCVIXI), prepared by catalytic hydrogenation of ü-£-tolyl- -o-isoglucosamine (XCVIl) in alkaline or neutral solution

(Weygand, 1940), with bromine in water gave o-mannose and D-fructose, identified ohromatographioally, and 2%6-dibromo- -j^-tolàidine ( C) • NH CHa 6:0 Oh' h o- 6 -h H-C-OH H-à-OH 6Hj.OH (XCVII). NH 6hi HO—6—ii H-Ç-ÛH H-Ç-OH 2 B i ^ 9*1 ffO-Ç-H liO-Ç-U il—C—OH fl—6—OH àHi.OH Br, D-Fruc tose and D-Mannose (XGVIII) (XCiX)

They showed 2:6-dibromo-II-j)-tolyl-u-mannamine (XCIX) to be an intermediate, and postulated a mechanism for the formation of u-mannose. These workers found it difficult to explain the formation of fructose; it was suggested that (XCIX) might be further oxidised to 2:6-dibromo-N-p-tolyl-p-isoglucosamine, which on hydrolysis would yield o-fructose. However, Weygand & Schaefer (193D were unable to identify any sugar products by treatment of N-p-tolyl-p-ieogluoosamine (XC/II) with bromine in water. Following neutralisation and evaporation of the

97.

aqueous solution obtained by bromine oxidation of N-^-tolyl- -D-mannamine these workers isolated the sugar products by

extraction of the residue with hot pyridine; it is surprising that o-glucose, formed by épimérisation of the fructose and/or the mannose by the action of the base, was not identified on the chromatogram.

It was considered by the present author that oxidation of N-p-tolyl-p-isogluoosamine in the manner described above would yield o-gluoosone; the inability of Weygand & Schaefer

(1931) to identify such a product of this oxidation may be explained on the grounds that the glucosone was converted into kojic acid, or a derivative thereof, by the extraction procedure with pyridine.

N-p-Tolyl-p-isoglucosamine was oxidised with bromine in water for 1 hour at room temperature. The almost colourless

solution obtained after décantation from the brown tar of

dibromo-j^-toluidine, removal of excess bromine with a stream of nitrogen, and neutralisation with sodium hydroxide gave the following reactions: it reduced Fehling's solution at room temperature, gave a negative result with Seliwanoff's reagent (thus showing the absence of fructose), did not regenerate the colour of Schiff's reagent, and gave a blue colour with

Benedict's arsenophosphotungstic acid reagent in the presence of alkali-cyanide; a small portion of the solution gave

D-gluoose phenylosazone on treatment with phenylhydrazine in acetic acid at room temperature; the presence of o-glucosone inljthe solution was confirmed by paper chromatographic analysis, employing the upper layer of a ii-butanol-aoetic aoid-water

(4:1:3) mixture as developing solvent and triphenyltetrazolium chloride ms identification reagent. The neutral aqueous

solution obtained as above was evaporated and the residue extr­ acted with absolute ethanol; the syrup obtained by evaporation of the ethanolic extract was treated with anhydrous acetone containing concentrated sulphuric acid with the production, in

low yield, of crystalline tri-O-isopropylidene o-glucosone hydrate, identical with the product of similar treatment of o-glucosone (see Part II, 3.2.2.). It was thus demonstrated beyond doubt that o-glucosone is formed by this reaction. Attempts to adapt the method to the preparation of the cryst­ alline isopropylldene derivative of o-glucosone on a large scale and in good yield were not successful, severe losses of the initial osone product being incurred during purification.

The following mechanism is proposed for the formation of o-glucosone (Cl) by bromine oxidation of N-p-tolyl-o-iso- glucosamine (XCVII), all compounds being represented in the open chain form:

1+