Biomedical Mass Spectrornetry 1974, l. 145 to 147

Chemical lonization and High Resolution Electron lmpact Mass Spectra of 1,6-Anhydro- 3,4- O-isopropylidene-P- D-talopyranose? DEREK HORTON, JON S. JEWELL, ERNST K. JUST and JOSEPH D. WANDER Department of Chemistry, The Ohio State University, Columbus, Ohio 43210, USA

RODGER L. FOLTZ Battelle Columbus Laboratories, Columbus, Ohio 43201, USA

(Receiced 1 3 Junuury 1974)

Abstract-The high resolution. electron impact mass spectrum of 1.6-anhydro-3.4-0-isopropyiidene-~-~-talo- pyranose. together with those of its derivatives having specific deuterium substitution at C-2, at C-3. at C-2 and C-3. and in the isopropylidene group. is considered in detail and compared with the ammonia and isobutane mediated chemical ionization spectra of these five compounds. From the elemental compositions of the fragment ion: and mass number shifts upon deuterium incorporation, the origin of the hydrogen atorns in each of the fragments is traced. and a detailed scheme of the main fragmentation pdthways is presected.

Introduction

THE ALDOSAN class of sugars, as exemplified by the 1.6-anhydrides of aldohexopyranoses, are of interest both for synthetic manipulation of sugars and for their characterization.2 They have a dioxabicyclo[3.2.1]- octane ring system and are formed in equilibrium with the aldohexoses when the latter are treated with dilute mineral acid, the extent of conversion being dependent on the stereochemistry of the aldohexose ~ s e d . ~ Maximal conversion is observed with (D or r)-idose and the product has the three hydroxyl groups disposed equatorially. whereas (D or L)-glucose under- goes conversion to the least extent among the eight stereoisomeric aldohexoses, to give the aldosan having ali three hydroxyl groups axialiy riented d.^,^ Being formed in conjunction with their parent aldohexoses in acid hydrolyzates of such natural products as poly- saccharides, glycoproteins, and complex glycosides, the aldosans are frequently encountered during struc- tural work on carbohydrates from biological sources in mixtures of sugars that require chromatographic separation and characterization. It is of interest, therefore, to elucidate in detail the mass spectral behavior of aldosans and their deriva ti ve^.^,^

In this laboratory the 1,6-anhydride of B-D-talo- pyranose has been of especial interest; it is the last stereoisomer of the class to have been synthesized’-’ and it accompanies D-talose (a sugar encountered rather infrequently in natural products)’ to the extent of about 12%) in acid hydrolyzates containing the sugar.3 1,6-Anhydro-B-~-talopyranose can react with acetone to give 5-membered ring acetals either at 0 - 2 and 0-3, or at 0 - 3 and 0 -4 . Although these isomers t Part ofa series Specific lsotropic Labeling of Sugars: Applications

in Mass-Spectral Anaiysis. For the previous report in this series. see Ref. 1.

CMe, O’ HO HO

1 .ó-Anhydro-D-o- talopyranose

2.3-Acetal

M e F B 0 ,

O

3.4-Acetal í 1)

are nominally equivalent, both being formed at an axial-equatoriai pair of hydroxyl substituents, the 2,3-acetaI6 is considerably more stable’ than the 3,4-acetal (1) (presumably because of the inductive effect of two oxygen atoms at C-1 in contrast to one at C-4) and is the principal product’ from direct acetonation reactions on the parent anhydro sugar.

In a previous report,s a detailed analysis of the electron impact, mass spectral fragmentation of the 2,3-acetal was achieved by use of high resolution techniques and a series of specifically deuterated derivatives. Cursory examination of the mass spectrum of the isomeric 3,4-acetal (1) revealed a surprisingiy different pattern of fragmentation behavior, and promp- ted the synthesis of specifically deuterated analogs, to permit the detailed analysis of its spectrum by high resolution techniques and by chemical ionization, which is the subject of this report.

As mass spectra are, in general, little influenced by changes of stereochemical configuration, the results obtained here for the derivative 1 of D-talose (a rela- tively uncommon sugar) should be equally applicable

145 O Heyden & Son Ltd., 1974

146 D. HORTON. J. S . JEWELL, E. K. JUST, J. D. WANDER A N D R. L. FOLTZ

for the common sugar D-galactose, which is readily converted" into the 2-epimer of 1. The data obtained by use of specifically deuterated derivatives allow the fate of the hydrogen atoms at C-2 and C-3 to be traced in detail, and should therefore form a useful basis for studies of metabolic interconversions conducted with appropriate specifically deuterated sugars as probes.

Experimental EQUIPMENT

Chemical ionization mass spectra of compounds 1 to 6 were obtained with a Finnigan quadrupole 101 5 m a s spectrometer equipped with a dual c.i.-e.i. source. The ion source was maintained at 200 "C with the ionizing gas pressure at about 0.5 Torr. The samples were introduced by means of a heated, direct-insertion probe. Under computer control (Systems Industries, Inc. 250 data system) the appropriate mass range was continuously scanned at approximately 5 s intervals. Al1 the mass spectral data were stored in the computer so that they could be displayed subsequently on a cathode-ray tube output device or by a digital plotter, either in the form of individual mass spectra, or as plots of selected m/e ion currents vs spectrum number. The filament was operated at an ionizing energy of lOOeV and an emission of 200pA. The ion repeller voltage was 8 V, the ion energy 9 V and the lens voltage - 75 V.

Electron impact mass spectra were recorded with A.E.I. MS-902 double focusing, high resolution mass spectrometers equipped with direct insertion probes, at an inlet temperature of 150 "C, an iotiizing potential of 70 eV and an accelerating potential of 8 kV. Mass spectra for data reduction were recorded directly on magnetic tape and converted into a digital signal before final processing on a Control Data 6400 com- puter, by using programs developed at Battelle Colum- bus Laboratories, Columbus, Ohio. The uncertainty in m a s numbers of fragments thus determined is no greater than 5 ppm.

MATERIALS 1,6-Anhydro-3,4-0-(isopropylidene-d6)-fi-~-

galactopyranose (7) 1,6-Anhydro-P-~-galactopyranose (250 mg) in ace-

tone-d, (6 mi) was stirred with p-toluenesulfonic acid (catalytic amount) for 12h at room temperature. Neutralization of the solution with solid sodium car- bonate and evaporation of the mixture, followed by extraction of the solid residue with dichloromethane (100 mi) gave, after evaporation of solvent, compound 7 as fine white needles; yield 280mg (90%), R , 0.25 (t.1.q silica gel, chloroform + ether, 3 : 1), m.p. 150 to 151 "C (Ref.lO, m.p. for undeuterated 7 150 to 151 "C).

1,6-Anhydro-3,4-O-(isopropylidene-d,)-fi-~-lyxo-hexo- pyranos-2-ulose (8) and its undeuterated analog (9)

In separate preparations 7 (250mg) and its un- deuterated analog (3.37 g) were oxidized to the cor-

responding 2-ketones with ruthenium tetraoxide by a procedure described previously7; yields of 8 and 9 were 210 mg (87 x) and 3.03 g (91 %), respectively. The 2-ketones 8 and 9, having physical constants in agree- ment with those reported.' were indistinguishable by t.1.c. (RF 0.38, silica gel, chloroform + ether, 3 : 1) and by mixed m.p. (92 to 93 "C, Ref. 7, m.p. 92 to 93 "C).

1,6-Anhydr0-3,4-0-isopropylidene-fi-~-lyxo- hexopyranose-2-ulose-3-d (10)

As described previously for the isomeric 4-ketone,' ketone 9 (1 g) was dissolved in deuterium oxide (1.5 mi) and its n.m.r. spectrum was recorded (100MHz. externa1 tetramethylsilane). A catalytic amount ( - 0.5 mg) of sodium deuteroxide was added and the spectrum was recorded at intervals, the sample being kept at - 25". After the elapse of 20 min, a one-proton multi- plet centered at z 5.65 had compietely disappeared with the concomitant appearance of an HOD signal of one- proton intensity. No further change was observed in the spectrum during 1 h at 25 "C.

The 3-deuterated ketone (10) was extracted into chloroform and the dried (sodium sulfate) extract was evaporated to give crystalline, chromatographically homogeneous 10 in almost quantitative yield. The product was indistinguishable from 9 by mixed m.p. and t.1.c.

1,6-Anhydro-3,4-0-isopropylidene-~-~-talopyranose (1) and its 2-deuterio (2), 3-deuterio (3), 2,n-di- deuterio (4) and 3,4-O-(isopropylidene-d,) ( 5 ) derivatives

Treatment of samples of the ketones 8 (200 mg) and 9 (500 mg) in separate ethanolic solutions (5 mi) with (a) excess sodium borohydride or (b) excess sodium borodeuteride (for 9 only), followed by neutralization (aqueous ammonium chloride), extraction into chloro- form, and drying (sodium sulfate), with subsequent evaporation of the extracts gave crystalline 1 (500 mg), 2 (450mg) and 5 (100mg), respectively. The products were recrystallized from ether + petroleum ether (b.p. 30 to 60°C). The same procedures were repeated, starting from the 3-deuterated ketone 10 (500mg), to give 3 (475mg) and 4 (500mg), respectively. The products 1 to 5 were indistinguishable from one another by m.p. (1 12 to 113 "C, Ref. 7 for undeuterated analog, m.p. 112 to 113 "C), mixed m.p., and t.1.c. mobility (R, 0.35, silica gel, chloroform + ether, 3 : 1).

Dissolution of 1 in deuterium oxide, followed by evaporation of the solvent, gave the 2-OD analog (6).

Results and discussion

Synthesis of specifically deuterated derivatives of 1,6-an- hydro-2,3-0-isopropylidene-/?-~-talopyranose (1)

To achieve full interpretation of the m a s spectrum of 1, it was necessary to have its analogs specifically C-deuterated at C-2 (2), C-3 (3), (2-2 and C-3 (4), and in the isopropylidene methyl groups (5). Each compound was obtained by a route starting from 1,6-anhydro-2,3-

SPECTRUM OF ~,~-ANHYDRO-~,~-~-ISOPROPYLIDENE-B-D-TALOPYRANOSE 147

O

H H H 1 6

t

CH,-O CH,-O CH,-O

M e 2 c W H / % M ~ c ~ H X M ~ C ~ :

O H H H 0 H D O H

H 2 1,6-Anhydro-2.3-0- I ,6-Anhydro-2,3-0-

kopropyhdene-D-D- isopropylidene-b-~- galactopyranose lyxo-hexopyranos-2-ulose

O H D H H

3 4 SCHEME 1

O-isopropylidene-P-D-galactopyranose (the 2-epimer of 1. readily available" by pyrolysis of lactose to afford a mixture containing 1,6-anhydro-~-~-galactopyranose as the only product having a cis-dio1 group that can react with acetone) and proceeding through oxidation at C-2 to 1,6-anhydro-3,4-0-isopropylidene-~-~-lyxo- hexopyranos-2-uIo~e.~ Reduction of the ketone with borodeuteride afforded the 2-iabeled alcohol (2) as this reduction proceeds ~tereospecifically~ to afford the talo alcohol (net inversion by the oxidation- reduction sequence). Base-catalyzed deuterium ex- change in the ketone takes place regiospecifically and stereospecifically at C-3 to give the 3-deuterio ketone, because the bridgehead hydrogen atom (H-1) cannot be removed by enolization (principle of Bredt's rule) and the thermodynamic stabiiity of the cis-acetal system assures that inversion at C-3 to give the less stable truns-acetal will not be favored. Reduction of the 3-deuterated ketone with borohydride gives the C-3 deuterated alcohol (3), and use of borodeuteride gives the 3,4-dideuterated alcohol (4). As direct acetonation of 1,6-anhydro-fl-~-talopyranose gives mainly the 2,3- acetal, the 3,4-acetal deuterated in the isopropylidene

group was prepared by acetonation of 1 ,ó-anhydro-P-~- galactopyranose with acetone-d,, with subsequent application of the oxidation-reduction sequence' at C-2 to afford the labeled talo derivative 5. The analog (6) of 1 O-deuterated at the 2-hydroxyl position was prepared by simple direct exchange with deuterium oxide.

By m a s spectrometry, compounds 2 to 5 behaved as products fully labeled at the positions indicated and showed no detectable labeling at other positions. The 'H n.m.r. spectrum of compound 1 and its deuterated analogs was not amenable to elementary interpretation on the basis of deuterium replacement (in contrast to the isomeric 2,3-acetal)" even with the use of para- magnetic shift reagents. ' Chemical ionization m a s spectral analysis of 1 and its

Data from these spectra are presented in Table 1. The spectra are exceedingly simple and the isobutane mediated spectra feature essentially three ions only, the protonated molecular ion [MH]', which is the base peak. accompanied by a weaker ion corresponding to

deuterated deriv2,tives (2 to 6)

148 D. HORTON, J. S. JEWELL, E. K . JUST, J . D. WANDER AND R. L. FOLTZ

loss of 58 Daltons from [MH]', and a still weaker peak attributable to loss of 58 + 18 Daltons from the [MH]' ion. Abstraction of C H , from the isopropylidene group

TABLE l . Intensities" ofmajor ionc in the c.i. (isobutane) m a s spectra of 1 to 6

mle I h 1 2 3 4 5 6"

220 100 - - - - ~~ [M + NH,]+ 209 ~ 100 - 205 - - - ~ 100 .- ~~

204 - - 100 100 -. - 100 203 52 100 - - - - - [MH]'

190 _ - - - - 5 - 8 ~~ 189 __ - - -

188 - - 7 6 -- 8 [M - CH,]' 187 8 8 - - -

162 2 - -. . .- - - [M + NH,]+ - Me,CO

147 146 ~- - 43 40 - 37 145 8 37 - - - 65 - [MH]' - Me,CO

129 - - - - 4 - - 128 -. ~ 6 6 - - 127 0.5 5 8 4 [MH]+ - Me,CO - H,O

a Expressed as percent of the base peak : in each case the base peak accounts for -5Ox of the ionization above m/e 100 after correction for isotopic peaks resulting from natural abundance.

38 - ~~

~~ - ~.

Arnrnonia c.i. 6 was prepared by uncatalyzed exchange of the hydroxyl proton

of 1 in D,O solution. Spontaneous reversal of this exchange with environmental protons appears to occur to sorne extent, either in handling or in the mass spectrometer, so that values presented for 6 were obtained by subtracting the spectrurn of 1 from the observed spectrurn in a proportion accounting for a 3:2 rnixture of 1 and 6.

by a gaseous hydrocarbon cation presumably leads to formation of a relatively weak peak at [M - 151, illustrated as a in Scheme 3.

The ammonia mediated spectrum of 1 shows each of the peaks already mentioned, together with an important ion resulting from ammonium ion capture [M + NH,]+ that supersedes [MH]' as the base peak in the spectrum; a minor ion resulting from loss of 58 Daltons from the new base peak is also evident.

The ammonia c.i. spectrum of 1 supports the ideal3 that this mode of ionization is the milder of the two in that intact capture ions account for >80% of the ion current above m/e 60, as compared with the corres- ponding - 50 % in the isobutane c.i. spectrum.

Reasons for the absence of ions resulting from capture of the bulky tert-butyl cation are not readily apparent, but the tendency of 1 to capture rather than deprotonate the [NH,]+ ion is an t i~ ipa ted '~ because of the lack of strongly basic functional groups in 1.

Under chemical ionization, the fragmentation of isopropylidene acetals of sugars appears to proceed virtually exclusively by loss of acetone.13 The data of Table 1 verify that the same mode of decomposition occurs for 1, which exhibits a reasonably prominent ion at m/e 145 (depicted as e in Scheme 3) formed by

loss of acetone from the [MH]' ion (the base peak), and indicate that the process of dehydration of m/e 145 [MH' - Me,CO] occurs only by loss of the 2-hydroxyl group plus either H-1 or H-4; deuterium labels in 5 disappear as expected upon loss of the elements of acetone, whereas 2, 3 and 4 show complete retention of the deuterium atom(s) at C-2 and C-3. Examination of the spectrum of compound 6, produced from 1 by conversion from the 2-OH into the 2-OD analog, shows exclusive loss of HOD from [MH' - Me,CO] (m/e 146), indicating that the 2-hydroxyl group is incorporated specifically into the water molecule that is eliminated. The resultant species most probably undergoes some rearrangement concomitant with or immediately subsequent to its formation, in order to alleviate the anticipated high strain that would de- stabilize such an unsaturated bicyclic ion, although there is at present no basis for specifying such a rearrangement.

Electron impact mass spectral analysis of 1 and its

The fragmentation pathways in the e.i. spectrum of 1 were elucidated by reference to the corresponding fragments in the data reduced, high resolution m a s spectra of the C-deuterio analogs 2 to 5.

Significant fragments formed from these five com- pounds after ionization (in a double focusing m a s spectrometer at source temperature of 150 "C) are recorded in Table 2, together with their intensities (relative to the base peak = 100) and the elemental composition of each fragment (expressed as the com- position of that ion in the m a s spectrum of 1). Table 2 is so arranged as to identify fragments undergoing shifts of m a s number because of isotopic incorporation.

In contrast to the 2,3-0-isopropylidene isomer,' the 3,4-acetal (1) and its deuterated derivatives (2 to 5 ) display weak but recognizable molecular ions [MI t A peak in the spectrum one mass unit higher arises through a competing (chemical) ionization process of proton capture, presumably from [MI+ ; this [MH]' peak is minute in spectra obtained by using a con- ventional spectrometer source, but is considerably more intense in spectra recorded on the instrument at Battelle Columbus Laboratories, where a special source of smaller interna1 dimensions (to favor the ion-molecule reactions necessary for chemical ionization) was em- ployed. Significant metastable peaks, which constitute an important and prominent feature in the spectrum of the 2,3-acetal and its deuterated analogs, occur less frequently and at much lower intensities in the present examples.

fragment ion (a) is favored by the ~ o m m o n ~ , ' ~ process of loss of a methyl radical from the isopropylidene group (Table 2 and Scheme 3) ; as with the 2,3-acetal, this ion confirms the molecular weight, although the relative proportions of this frag- ment and the acetylium ion [m/e 43(46)] do not appear to reflect a systematic secondary isotope effect of the

deuterated derivatives 2 to 5

The abundant C8H

SPECTRUM OF ~,~-ANHYDRO-~,~-~-ISOPROPYLIDENE-~-D-TALOPYJWNOSE 149

TABLE 2. lntensities of selected fragments in the electron-impact m a s spectra of compounds 1 and 2 to 5, expressed as percent of the respective base peaks

Compound Elemental m/e 1 2 3 4 5 Composition

209 - -- - 9.0 C9H905D6 205 - - - 8.0 - C,HI,O,D2 204 - 13.2 19.8 - - C9H

203 9.7 - - - - C9H 15'5

208 - - - - 0.3 C9H,05D6 204 - - - 0.5 - C9H1205D2

203 - 0.7 0.3 - - C9H1305D

202 0.3 - - - - C9H 14'5

190 -- - - - 74.5 C,HSO,D, 189 - - - 49.4 - C8H905D2

188 - 81.1 82.3 - - CüH1005D

187 58.7 - - - - CSH1105

191 - - - - 1.8 C9H704D6

186 -. 1.0 1.8 -- -

179 - - - - 0.4 C6H704D6

187 - - - 1.0 - C9H 1 1°4D2

C9H1

185 1.0 - - ~- - C9H1304

174 - - 0.3 0.2 - C6H 1

173 0.6 0.4 - - - C8H1304

161 - - - 0.1 - C7H904D2

160 - 0.1 0.1 - - C7H 10°4D

159 0.2 - - - - C7H1104

158 - - - 0.2 - C6H1003D2

157 - 0.5 0.3 - - C 8 H l I O P 156 0.6 - - -~ - C8H1203 147 - - - 7.7 - C6H704D2

146 - 15.2 17.5 - 2.4 C6H804D 145 9.8 - - - 10.0 C6H90, 149 - - - - 7.5 C7H503D6 143 6.8 9.3 11.0 7.0 - C7Hl103 145 - -- - 1.2 - ' 6 5

144 - 2.1 1.8 - - C6H604D

143 1.1 - - - 1.0 C6H,04 144 - - - - 0.2 C7H603DJ

C7H703D2 143 - - - - - 142 - 0.4 0.3 - - CTHBO3D 141 0.4 - - - - C7H903

139 2.0 2.1 - - - CSHl 1 0 2

145 - - - - 3.5 C8H50,D6 140 - - 3.3 2.0 - C,H,OO2D

141 - - - 0.4 - C7HSO3D2 140 - 0.3 0.2 - - C7H603D

139 0.2 - - - 0.4 C7H703 133 - - - - 1.5 C7H50,D6 129 - - - 0.2 - C7H902D2

128 - 1.0 0.9 - - C7Hi002D

127 1.2 ~

129 - - - 6.8 - C6H503D2

128 - 11.9 13.7 2.7 - C6H603D

127 8.6 - 3.9 - 10.7 C6H703

C7H1 lo, .. - -

129 - - - - 0.2 C6H,03D3 128 - - - 2.6 - C,H,O,Dz 127 - 3.9 3.9 - - C6H503D

Elemental Compound m/e 1 2 3 4 5 composition

126 1.4 - - - 0.9 C6H603 117 - - - 0.8 - C5H503D2

116 - 2.3 3.2 1.4 - C5H603D 115 2.8 2.4 1.9 - 3.5 C5H703 119 - - - - 0.2 C6H,02D6 115 - - - 0.3 - C6H702D2

114 - 0.7 0.5 - - C6H602D

113 1.1 - - - -

110 - 2.9 - 1.9 - C6H402D

C6H902

109 2.4 - 3.2 - 3.6 C6H502 107 - - - - 5.8 C5H30,D6 103 - - - 3.3 C5H702DZ 102 -

101 3.4 2.8 3.9 2.3 2.9 C 4 H 5 0 3 106 - - - - 6.9 C5H202D6

C5H602D 6.6 6.1 - - 101 5.2 ~ - - - C5H902

101 - - 10.6 11.5 - C 5 H 7 0 2 D

100 3.4 11.7 - - - 101 - ~ 11.5 - C5H50.?D2

100 - 11.7 15.6 10.0 - C5H602D

C,H,O,

99 11.7 - 12.7 - 10.5 C5H702 103 - - - - 1.1 C6H30D6 97 5.7 6.2 7.7 4.5 - C6H90

9 9 - - - 5.4 - C 5 H 3O2 D2

98 - 15.2 8.9 6.8 - C5H40,D 97 11.9 - 10.6 - 12.0 C ~ H ~ O Z 8 8 - - - - 10.5 C4H202Dj 86 - - 26.3 16.7 - C4H402D

- 21.3 C4H5O2 85 26.0 31.0 - 8 5 - - - 8.8" - C4H02D2

84 - 1.4 1.5 - - C4H202D

83 1.4 - - - 1.3 C4H,0,

82 - 33.2 20.0 - - C 5 H 4 0 D 8 3 - - - 11.1 - C5H30D2

81 25.4 - - - 30.4 C5H5O 7 5 - - - 5.4 - C3H302D2

74 - 12.3 10.1 ~ - C3H402D

73 14.6 - - - 13.6 C3H5OZ 1 3 - - - 2.7 2.7 C4H50D2

71 6.0 - - - 5.0 C4H70 C4H60D 72 - 5.7 4.6 - -

7 3 - - - 8.2 - C3HO2D2 C3H202D 72 - 17.2 12.9 - -

71 15.7 ~ 16.5 C3H302 71 - - 17.6 12.2 - C4H50D

C4H40D - 20.5 C4H60

- 43.1 C4H50

70 15.4 24.8 - 10 - - 40.0 25.3 - 69 34.5 43.1 - 6 5 - - - - 85.8 C3HODb 59 75.0 i00.obi00.ob 68.5 - C3H7O

- iOaOb C,D,O 4 6 - - - 43 iOO.Ob 75.0 63.0 100.Ob - ' Z H 3 0

a A second ion [C4H4D0,]+ contributes to this value. The base p&k represents 1 1 "', (l), 12.5% (2), 12% (3), 15:/ú (4) and lo:( (5). respectively, of the net ion current between m/e 43 and 300.

type noted' for the isomeric 2,3-acetal. The next most massive species formed by decomposition of [MI f

hydroxyl proton to C-3.t Fragment a decomposes further by loss of ketene to yield the C6H,0, ion already

is the low intensity C,H, 30, ion (b), which evidently arises by loSS of .OH from [MI t, A third high maSS

'pecies (C8H13043 '1, Of 'Ow intensity3 may be formed by scission of the C-1 -C-2 and C-2-C-3

t The structures given for the ions depicted in the Schemes in this paper are consistent with the data of the molecular formulae and the retention of isotopic labels. Alternative, isomeric structures of the Same moiecuiar formula are feasible and are not excluded, especially

bonds of [MI with concomitant migration of the for the smaller ions.

150 D. HORTON, J. S. JEWELL, E. K. JUST. J. D. WANDER AND R. L. FOLTZ

+ oo OH HO

e

[m/e 145(1), 146(2. 3.5). C6H904

147(4)1

-CHICO T g ? j =

Me a

[m/e 187(1), 188(2,3), C*Hl ,O,

189(4). 190(5)1

Me Me Me

Me c b

C9H 13'4

[m/e 185(1), 186(2,3). l87(4). 19 1(5)]

C8H1304

79(5)1 [m/e 173(1, 2). 174(3, 4).

-C,H,O 1

Me, .COH4 1

SCHEME 3

noted in the c.i. spectra, although direct loss of acetone from the [MH]+ ion presumably contributes sub- stantially to the abundance of this fragment. It may be formulated as e or as the corresponding 4-hydroxy C-3 cation. The C,H,,O, ion may arise from a. by scission of the C-1-C-2, C-1-0-6 and C-5-0-5 bonds, with concomitant migration of H-1 to 0-6, to afford f. Further decomposition of b proceeds by loss of HC-2 and HC-3 with 0 - 3 as the elements of ketene to give C,H, formulated as g ; subsequent elimina- tion of H-4 and C-1 (with 0-5 and 0 - 6 attached) as formic acid generates the C,H,O fragment ( h ) ; a weak metastable ion at m/e 68.5 (calc. 68.54) signals the loss from g of CO, to form a C,H , ion and a second weak metastable ion at m/e 24.3 (calc. 24.34) corres- ponds to separation of a protonated acetone molecule from g. Initial loss of H-4 as a component of formic acid from b leads to C,H , whereas a second minor fragment at m/e 139 (C,H,O,) appears to arise by less elementary processes in which both isopropylidene methyl groups are lost, together with two oxygen atoms and an unlabeled hydrogen atom. Loss of the elements of acetone plus acetaldehyde from b to form a C5H loO, ion is indicated by a weak metastable ion at m/e 37.2 (calc. 37.24). Elimination of acetic acid from a (Scheme 4) gives C,H,03 (i), which is verified by a weak metast-

able ion at m/e 86.2 (calc. 86.25); i apparently suffers subsequent dehydration to C,H,O, with loss of H-3.

subsequent to rearrange- ment, by loss of a formylhydroxymethyl radical may be invoked to rationalize C,H ,O, as a vinyldioxolanium ion ( j ) , at the same m a s number as i.

One C,H,03 ion (k) can arise by loss of acetone from c, or by synchronous loss of acetone plus a formyl radical from [M]i , as illustrated in Scheme 5, whereas an approximately equally abundant isomer (k') may result from decarbonylation of the intermediate d ; only H-3 is retained in the former, whereas both H-2 and H-3 are retained in the latter. Subsequent loss of the elements of formic acid from k to form a C,H,O ion retaining H-3 specifically is indicated by a weak metastable ion (m* 41.3, calc. 41.40); this C,H,O ion may be used to distinguish the presence and extent of labelling of H-3 and, by difference, of H-2.

Loss of an hydroxyl radical from d or. alternatively. concerted loss of acetone hydrate from [MI" would produce the highly strained but resonance-delocalized fragment (1). A less abundant isomer (í'), in which one of the isopropylidene methyl groups is retained, can be formed from a by a series (Scheme 6) of processes in which hydrogen atoms from 0 - 2 and C-4 migrate to 0 - 5 and C-6, the C-1-C-2, C-5-C-6, and O-5-C-5

Decomposition of [MI

SPECTRUM OF ~,~-ANHYDRO-~,~-~-ISOPROPYLIDENE-~-D-TALOPYRANOSE 151

r H 1 l Me O

Me Me O [MI? 4

Me j

C , H , , 0 2 [m,k 127(1), 128(2,3).

129(4). 133(5)]

SCHEME 4

bonds break, and an O-2-C-5 bond forms, with loss of the methyl hemiacetal of a formyl radical.

Decomposition of b by loss of an even-electron dioxolenyl species yields C6H902, presumably the dioxenylium ion (m) ; the formation process illustrated (Scheme 6) is similar to that proposed’ for formation of the same ion from the 2,3-isomer of 1, and a weak metastable ion (m* 68.9, calc. 69.02) accords with this formulation. Sequential losses of formic acid and of the methyl radical from [MI? give the ions C,H,,03 (n) and C,H90, (o), respectively. These three ions were identified’ as minor contributors in the decomposition of the 2,3-acetal.

Although the C,H,O fragment appears to be iso- merically unique, in that it retains both H-2 and H-3

completely, the isotope distribution patterns in the C,H,O, (hydroxypyrilium) ion, and in the presumably reiated ions C,H,03 ( i , i’) and C,H,02, suggest that competing pathways act to generate isomeric forms of these three ions. Decomposition (Scheme 7) of i by loss of C-6 as formic acid rationalizes the formation of C,H,O ( p ) ; associated metastable evidence indicates that the decomposition steps a --+ i + p proceed both through a fairly stable intermediate i (m* 51.8, calc. 51.66) and also essentially directly from a to p (m* 35.0, calc. 35.09).

Scission of the C-1-C-2, C-3-C-4 and O-4-iso- propylidene bonds of [MI? with retention of the charge in either of the fragments would produce the isobaric C,H,O, and CSH9O2 fragments (4 ) and (y)

OH k

[m/e 1131, 2. 5).

[MI: Me

CSH,O3

1 16(3. 4)]

SCHEME 5

152 D. HORTON, J. S. JEWELL, E. K. JUST, J. D. WANDER AND R. L. FOLTZ

Me O‘H

a I’

[m/e 1201) 127(2, 3). C,H,O,

128(4), 129(5)]

Me Me b m

C6H902

1 134). 1 19(5)] [m/e 113(1), 114(2,3),

[MI f n O

C8H1203 C7H903

158(4), 162(5)] 143(4), 144(5)] [m/e 156(1), 157(2.3), [m/e 141(1), 142(2. 3),

SCHEME 6

c$ -HC*02H @ 2 -HC60 H &ej - AcOH m’35.0

- m‘S1.8

Me

(Scheme 8), whereas cleavage at C-2-C-3 and C-4-C-5 would liberate CsH80, (formulated as the dioxolenium radical cation (s), which may further decompose by loss of a methyl radical to generate the abundant C,HSO2 cation (t). The formation of these four frag- ments appears exactly analogous to their genesis in the decomposition’ of the 2,3-acetal and the decomposi-

tion of s -+ t is verified by a metastable ion at m/e 72.2 (calc. 72.25).

The ion C6HSO2 (m/e 109 in 1) retains H-2 to the total exclusion of H-3, whereas the ions C,H,O (m/e 70) and C,H50 (m/e 69) feature specific retention of H-3 to the total exclusion of H-2. Accordingly, parallel examination of these three ions for m a s number shifts

SPECTRUM O F ~.~-ANHYDRO-~,~-~-ISOPROPYLIDENE-~-D-TALOPYRANOSE 153

J$zJL O- F{ and O OH

Me Me M e 4 u Me J

[MI: 4 r C A O , C5H902

[m/e IOi] [mie 101(1), 102(2,3), 103(4), 107(5)]

SCHEME 8

offers a direct means of assaying the extent of deu- terium replacement at H-2 and H-3, provided that substitution is limited to these two positions. This may be determined by reference to the C,H,O, ion (m/e lOO), which contains H-2 and H-3 as the only protons remaining from the original skeleton, and to the C8H, ion (u, m/e 187), which retains al1 of the original skeletal hydrogen (or deuterium) atoms. Location of isotopic labels replacing atoms other than H-2 or H-3 would require positive identification of fragments in which the other atoms are specifically retained or eliminated.

The foregoing data for 1 clearly illustrate the analyti- cal value of using both the c.i. and e.i. modes in con- junction to afford complementary data. The c.i. mode gives the molecular weight and proves an excellent check of homogeneity, as impurities are readily recognized;I5 identification of 1 as a component in a mixture of compounds could readily be effected, as could the determination of the presence (and relative amounts) of isotopically substituted forms of the molecule. In contrast, the e.i. mode is of limited use with mixtures and does not provide the most sensitive and direct method for determining molecular weight, but it does generate a range of observable fragmenta- tions that can be used to establish the positions in the molecule at which isotopic substitution has been effected. Although the data presented here refer speci- fically to compound 1, these general principles should be pertinent in the broader context of analysis of other sugars and their derivatives, including isotopically sub- stituted forms, obtained during structural work on biological molecules and in transformations of sugars by chemical or enzyme-catalyzed processes.

ACKNOWLEDGMENT The authors thank C. R. Weisenberger of this Department for

recording some of the m a s spectra employed in this study. This work

was supported, in part, by a grant from the National lnstitutes of Health, Public Health Service, Department of Health, Education and Welfare, Bethesda, Maryland 20014; Grant No. GM11976 (The Ohio State University Research Foundation Project 1820) and, for the data-reduced spectra, by NIH Contract No. 69-2226 to Battelle Columbus Laboratories.

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