Multisite modification of neomycin B: Combined Mitsunobu and click chemistry approach

Multisite Modification of Neomycin B: Combined Mitsunobu and

Click Chemistry Approach

Sabina Quader,^†Sue E. Boyd,^†Ian D. Jenkins,*^,§and Todd A. Houston*^,‡

Eskitis Institute and Natural Product DiscoVery, Griffith UniVersity, Nathan, QLD 4111, Australia, and

Institute for Glycomics, Griffith UniVersity, Gold Coast, QLD 9726, Australia

i.jenkins@griffith.edu.au; t.houston@griffith.edu.au

ReceiVed October 10, 2006

The aminoglycoside antibiotic neomycin B has been converted into several novel building blocks that

can be used for the specific modification of three of the four ring systems. Under carefully controlled

conditions, the Mitsunobu reaction can be used to selectively dehydrate the ido ring to give the talo

epoxide. Subsequently however, under more forcing conditions, the 2-deoxy streptamine ring undergoes

Mitsunobu dehydration to give an aziridine. An unusual remote neighboring group effect was observed.

When the primary hydroxyl of the ribose ring was blocked, aziridine formation on the deoxystreptamine

ring did not occur. Both the epoxide and epoxide-aziridine neomycin building blocks can be ring-opened

with azide and subjected to “click” type chemistry with terminal alkynes to generate a series of new

neomycin analogues. These reactions can all be carried out without recourse to O-protecting groups. A

detailed conformational analysis by NMR revealed some unexpected conformer preferences in these

systems.

Introduction

impedes mRNA translation, results in miscoding, and leads

ultimately to bacterial cell death.²Compounds in this class can

interact with a variety of other biologically relevant RNA

sequences including group I introns,³hammerhead ribozymes,⁴

and the hepatitis delta virus (HDV) ribozyme.⁵Recently, the

aminoglycoside antibiotic neomycin B, 1 (Figure 1), has also

been found to be the most potent inhibitor of the proteolytic

activity of anthrax lethal factor (LF).⁶Despite the broad-

spectrum efficacy of these natural amino sugars, the use of these

Aminoglycoside antibiotics are widely used for the treatment

of a variety of infections including tuberculosis and pneumonia

in particular. They exert their antibacterial activities by binding

to RNA sequences in the bacterial ribosome,¹and this binding

^†Eskitis Institute.

^§Natural Product Discovery.

^‡Institute for Glycomics.

(1) (a) Tanaka, N. In Handbook of Experimental Pharmacology;

Umezawa, H., Hooper, I. R., Eds.; Springer- Verlag: New York, 1982; pp

221-266. (b) Cundlife, E. In In the Ribosome; Hill, W. E. et al, Eds.;

American Society of Microbiology: Washington, DC, 1990; pp 479-490.

(c) Noller, H. F. Annu. ReV. Biochem. 1991, 60, 191-227.

(2) (a) Moazed, D.; Noller, H. F. Nature 1987, 327, 389-394. (b) Puhroti,

P.; Stern, S. Nature 1994, 370, 659-662. (c) Recht, M. I.; Fourmy, D.;

Blanchard, S. C.; Dahlquist, K. D.; Puglisi, J. D. J. Mol. Biol. 1996, 262,

421-436. (d) Miyaguchi, H.; Narita, H.; Sakamoto, K.; Yokoyama, S.

Nucleic Acids Res. 1996, 24, 3700-3706.

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(6) (a) Lee, L. V.; Bower, K. E.; Liang, F.-S.; Shi, J.; Wu, D.; Suchek,

S. J.; Vogt, P. K.; Wong, C.-H. J. Am. Chem. Soc. 2004, 126, 4774-5. (b)

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Published on Web 02/14/2007

1962

J. Org. Chem. 2007, 72, 1962-1979

Multisite Modification of Neomycin B

selective chemistry elsewhere in the molecule, and this has been

exploited for modification of the ido and streptamine ring

systems. We have developed highly selective chemistry on this

natural product that allows for incorporation of an azide

functional group in each of three rings of neomycin and further

transformation via copper-catalyzed coupling reactions with

hydrophobic alkynes. This chemistry precludes the need for

O-protecting groups and thus allows for higher synthetic

throughput.

Our initial aim was the introduction of hydrophobic esters

that might serve as prodrugs to improve the parent drug’s

effectiveness at crossing the waxy, mycolate-rich barrier that

envelops Mycobacterium tuberculosis.¹¹Although the ami-

noglycoside streptomycin was the first effective drug in the

treatment of tuberculosis, drug resistance to this class of

compounds was observed quite early after its introduction and

is a growing problem today.¹²It has recently been demonstrated

that mycobacteria enter the macrophage by way of a cholesterol-

dependent mechanism.¹³Membrane cholesterol depletion re-

duced macrophage uptake of M. tuberculosis by 85%. Normally,

after phagocytosis mycobacteria actively recruit and retain

tryptophan aspartate-containing coat protein (TACO) and this

prevents delivery to lysosomes, thus protecting the pathogen

from degradation. The TACO associates with the phagosome

in a cholesterol-dependent manner, and it is known that

mycobacteria display a high binding capacity for cholesterol

itself. By modifying antitubercular drugs with mimics of this

steroid,¹⁴it may be possible to hijack this binding mechanism

as a route for targeted drug delivery.¹⁵

FIGURE 1. Conventional numbering scheme for neomycin B, 1, and

the numbering scheme adopted here for illustrative clarity (inset).

drugs is restricted due to their toxicity, poor oral bioavailability,

and poor cell permeability. Consequently, semisynthetic ana-

logues of this class of compounds are attractive targets for

medicinal chemists,⁷as modified compounds of this type may

demonstrate increased effectiveness in clinical applications.

Although there are many examples of direct synthetic modifica-

tion of aminoglycosides, these often require multiple protection/

deprotection strategies using a variety of O-protecting groups,

as well as N-protecting groups, to both improve the organic

solubility of these hydrophilic compounds and to allow the

selectiVe manipulation of the numerous amino and hydroxy

substituents. Furthermore, even though naturally occurring

neomycin B has several potential points of attachment for the

addition of substituents to attenuate the properties of the drug,

little effort has been directed toward modification of all four

residues of the tetrasaccharide scaffold. Indeed, although many

structural analogues of this antibiotic have been synthesized over

the past decade, the majority of these cases involve modification

at the ribose moiety, by exploiting the differential reactivity of

the single primary hydroxyl group.

“Near perfect” reactions are required to maximize the

synthetic efficiency of selective modifications of complex

molecules such as the amino-glycoside antibiotics. The develop-

ment of the concept of “click chemistry” by Sharpless has

provided the organic chemist with a first generation toolbox of

such reactions.⁸Two identified by Sharpless⁹involve azides:

nucleophilic opening of epoxides and coupling reactions with

alkynes. Here, we describe the use of this chemistry, in

conjunction with Mitsunobu dehydration reactions, for the rapid,

highly selective modification of the aminoglycoside antibiotic

neomycin B in three different ring systems. Having several

potential points of attachment would allow addition of substit-

uents to improve the nonspecific physicochemical properties

of the drug without destroying the specific biological activity.

Initially we sought a more direct and specific functionalization

of the singular primary alcohol found in many members of this

family through Mitsunobu esterification, previously demon-

strated to be successful in this regard on disaccharides such as

trehalose.¹⁰However, the Mitsunobu reaction has resulted in

Results and Discussion

Modification of the ido Ring. It has been reported by Fourrey

et al. in two separate publications^16,17that under the conditions

of the Mitsunobu reaction the ido ring of hexa-N-Boc-protected

neomycin B forms a tricyclic aziridine-azetidine structure. We

have shown¹⁸that this structure is incorrect, and that an epoxide

is formed, as expected for the ido conformer with trans-diaxial

alcohols. Thus, when 2 was treated with 2 equiv of triph-

enylphosphine (TPP) and diisopropyl azodicarboxylate (DIAD),

the epoxide derivative of neomycin B, 3, was formed in 40%

(10) (a) Bottle, S.; Jenkins, I. D. Chem. Commun. 1984, 385. (b) Jenkins,

I. D.; Goren, M. B. Chem. Phys. Lipids 1986, 41, 225-235.

(11) For examples of lipophilic antitubercular compounds, see: (a)

Ballell, L.; Field, R. A.; Duncan, K.; Young, R. J. Antimicrob. Agents

Chemother. 2005, 49, 2153-2163. (b) Jones, P. B.; Parrish, N. M.; Houston,

T. A.; Stapon, A.; Bansal, N. P.; Dick, J. D.; Townsend,C. A. J. Med. Chem.

2000, 43, 3304-3314. (c) Parrish, N. M.; Houston, T.; Jones, P. B.;

Townsend, C.; Dick, J. D. Antimicrob. Agents Chemother. 2001, 45, 1143-

1150.

(12) Smith, C. V.; Sharma, V.; Sacchettini, J. C.; Tuberculosis 2004,

84, 45-55.

(13) Gatfield, J.; Pieters, J. Science (Washington, D. C.) 2000, 288,

1647-1650.

(14) For a cholesterol-derived aminoglycoside designed for gene trans-

fection, see: Belmont, P.; Aissaoui, A.; Hauchecorne, M.; Oudrhiri, N.;

Petit, L.; Vigneron, J.-P.; Lehn, J.-M.; Lehn, P. J. Gene Med. 2002, 4, 517-

526.

(15) Alternatively we are interested in designing boronates to target the

mannose-rich exterior of M. tuberculosis, see: (a) Gray, C. W., Jr.; Walker,

B. T.; Foley, R.; Houston, T. A. Tetrahedron Lett. 2003, 44, 3309-3312.

(b) Johnson, L. L., Jr.; Houston, T. A. Tetrahedron Lett. 2002, 43, 8905-

8909.

(16) Hernandez Linares, A.; Fourmy, D.; Fourrey, J.-L.; Loukaci, A. Org.

Biomol. Chem. 2005, 3, 2064-2066.

(17) Hernandez Linares, A.; Fourmy, D.; Fourrey, J.-L.; Loukaci, A.

Synth. Commun. 2006, 36, 487-493.

(7) For a review of methodologies in syntheses of aminoglycoside

antiobiotics, see: Haddad, J.; Liu, M.-Z.; Mobashery, S. In Glycochemis-

try: Principles, Synthesis and Applications; Wang, P. G., Bertozzi, C. R.,

Eds.; Marcel Dekker: New York, 2001; pp 353-424.

(8) Kolb, H. C.; Finn, M. G.; Sharpless, K. B. Angew. Chem., Int. Ed.

2001, 40, 2004-2021.

(9) Kolb, H. C.; Sharpless, K. B. Drug DiscoVery Today 2003, 8, 1128-

1137.

J. Org. Chem, Vol. 72, No. 6, 2007 1963

Quader et al.

SCHEME 1^a

^aReagents and conditions: (a) TPP (2 × 2 equiv), DIAD (2 × 2 equiv), toluene, rt, N₂, 18 h; (b) TPP (4 equiv), DIAD (4 equiv), toluene, rt, N₂, 18 h;

(c) 4-NO₂C₆COOH (12 equiv), TPP (10 equiv), DIAD (10 equiv), toluene, rt, N₂, 18 h; (d) MeOH/28% NH_3(aq), 8 h, rt; (e) NaN₃, rt, DMF, 2 d; (f)

R′COO-CH₂C′′CH (2 equiv), t-BuOH/H₂O (1:1), Cu/CuSO₄, rt, 18 h.

yield.^18aEpoxide 3 represents an attractive target for subsequent

selective modification of the neomycin B framework at the ido

moiety (Scheme 1). However, under our initial synthetic

conditions almost 50% of starting material, 2, was recovered

from the reaction mixture unconverted. In an effort to increase

the conversion of the starting material and thus to increase the

yield of the epoxide 3, the reaction was repeated using an excess

of the two reagents. Accordingly, compound 2 was treated with

4 equiv of TPP and DIAD at room temperature for 12 h in

toluene. However, although the net conversion of the starting

material 2 did improve, the yield of the epoxide 3 did not. After

column chromatographic separation, the monoanhydro com-

pound 3 was found to have formed in only in 33% yield.

Interestingly, a bis-anhydro neomycin derivative, 4, (HRMS

[M + H⁺] ) 1179.60897) was obtained as the major product

(45%). The same bis-anhydro species was found to be present

as a minor product in the initial reaction. The structural

characterization and chemistry of 4 is discussed further in the

succeeding sections. An increased yield of the mono-anhydro

species 3 (62%) was obtained when 4 equiv of DIAD and TPP

was added in two portions of 2 equiv each, with a 12 h interval

between additions. However, optimal yields of 3 are obtained

if an alternate, two-step, procedure is employed. Treatment of

2 with 12 equiv of 4-nitrobenzoic acid, under Mitsunobu

conditions (10 equiv of TPP and DIAD), affords the 5_III-

substituted 4-nitrobenzoate ester derivative 5 in high yield

(85%). Esterification of the primary hydroxyl of the ribose

residue suppresses the formation of the side products (vide infra).

Ester 5 was then readily converted to 3 on treatment with

methanolic aqueous ammonia solution (2:1, MeOH/ 28%NH_3(aq)

)

by simply stirring the suspension at room temperature (8 h,

quantitative). Per-acetylation of the mono-anhydro product using

acetic anhydride and pyridine afforded penta-O-acetyl mono-

anhydro neomycin B, 3-penta-O-acetate.

The structure of 3-penta-O-acetate was probed using high-

temperature ¹H NMR spectroscopy.^18aThe formation and

stereochemistry of the oxirane ring in 3 can be inferred from

the structure of the product of the ring opening reaction of 3

with azide ion. Treatment of the mono-anhydro product, 3 with

NaN₃in DMF at room temperature for 2 days furnished an azido

neomycin 6 (Scheme 1) as a single product (85% yield after

purification by column chromatography). Alternatively, 6 could

be obtained directly from the ester 5, on treatment with NaN₃.¹⁹

The regioselective ring opening of the talo epoxide in 3 and 5

at C_3IVby azide ion to give the trans-diaxial product (ido

configuration) follows from the Fu¨rst-Plattner rule and from

consideration of the relative energies of the two possible epoxide

(18) (a) Quader, S.; Boyd, S. E.; Jenkins, I. D.; Houston, T. A. Org.

Biomol. Chem. 2006, 4, 36-37. (b) The errors in the structures initially

reported by Fourrey et al. in ref 16 are noted in footnote 11 of ref 18a

(which formally corrects the aziridine/azetidine structure presented in ref

16). The subsequent errors in the similar structures reported in ref 17 are

acknowledged by Fourrey et al. in the unreferenced footnote, footnote [15-

(PDF version)/18(HTML version)], which renounces the aziridine/azetidine

structures presented in the main body of that article. Professor Fourrey has

advised us in a personal communication that after a re-examination of their

NMR data, they were unable to observe the HMBC correlations between

the respective Boc-carbonyl carbons and H-3′′′ and H-4′′′ protons, forwarded

initially as diagnostic evidence for the aziridine/azetidine structure described

in ref 16.

(19) Go´mez-Vidal, J. A.; Forrester, M. T.; Silverman, B. Org. Lett. 2001,

3, 2477-2479.

1964 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

SCHEME 2

azido derivative, 6, some 9.2 ppm to high field of the

corresponding resonance in the parent compound, 2 (70.0 ppm).

These shifts are in accord with those provided in previous reports

of azidosaccharide species.²³The stereochemistry of the azide

addition product, 6, was confirmed using selective 1D NOE

experiments conducted on 6-hexa-O-acetate. Figure 3a shows

1

an expansion of the H NMR spectrum of 6-hexa-O-acetate,

focusing on the region containing the H_3IV, H_5IV, and H_2IV

resonances. Selective saturation of the near co-incident H_4IVand

H_1IVresonances (δ 5.13 ppm) results in NOE enhancement of

the H_3IV, H_5IV, and H_2IVresonances, as expected.²⁵Selective

irradiation (δ 6.43 ppm) of the overlapping H_(2)NH(IV)and

H_(2)NH(I)resonances results in significant enhancement of the

H_3IVand H_2IVresonances, establishing the proximity of the H_3IV

and H_(2)NH(IV)nuclei.²⁶This places the H_3IVproton on the top

face of the ido ring (IV), i.e. in the equatorial position,

confirming axial attachment of the azido moiety and thus, by

inference, the talo configuration of the monoanhydro precursor,

3.^18aThe singlet resonance expected for the aryl proton of the

newly formed triazole unit is in evidence in the ¹H NMR

spectrum (Figure 2c) of the click product, 8-hexa-O-acetate,

(8.40 ppm), as is the deshielded broad singlet observed for the

methylene protons adjacent to the ester linkage (5.67 ppm). The

resonances attributable to H_3IVand H_4IVboth show distinctive

downfield shifts (5.41 and 5.75 ppm) upon conversion of the

azide substituent to the triazole ring (cf. Figure 2, b and c).

The ¹³C NMR chemical shifts of C_3IVand C_4IVare less sensitive

to the conversion of the azide substituent to the triazole unit,

however. In 8-hexa-O-acetate, C_3IVand C_4IVare observed at

61.6 and 69.9 ppm, respectively, both within 2 ppm of the

corresponding resonant frequencies of these carbons in the per-

acetylated azido precursor, 6-hexa-O-acetate.²⁷

The success of the model alkyne-azide coupling reaction

encouraged us to adopt this click approach as our general

method of choice for lipophilic modification of the ido residue

of the neomycin B core. The click synthons propargyl laurate,9,

and a propargyl ester of a cholesterol derivative, 10, illustrate

the utility of the procedure for increasing the hydrophobicity

and biological recognition and transport properties of the

aminoglycoside. Mycobacterial interaction with membrane-

bound cholesterol must involve some sort of recognition of the

A-ring, as this portion of the steroid will be oriented toward

conformations (Scheme 2).²⁰The ^OH₁conformation (3-A) would

be expected to be lower in energy than the H_Oconformation

1

(3-B), as there is a severe (pseudo) 1,3-diaxial interaction

between the C₆NHBoc group and the ribose moiety at C₁in

3-B. Conformation 3-A may also be stabilized by a hydrogen

bonding interaction between the C₂NH and the ring oxygen.

Similar energetic considerations are likely to be reflected in the

relative energies of the transition states leading from 3-A and

3-B. Additionally, attack on conformation 3-A is likely to be

further favored over 3-B as the transition state for attack by

azide ion on conformation 3-B will be subject to an additional

dipolar repulsion (field effect) between the attacking nucleophile

and the ring oxygen atom.

The regiospecific opening of the epoxide 3 by azide affords

a new functionality in the ido ring with potential for further

facile modification using a click chemistry approach; in this

case, the copper-catalyzed coupling of appropriately substituted

acetylenes to the azide substituent. Such a utilitarian procedure

offers wide scope for the attachment of lipophilic groups to the

neomycin B core. In order to examine the feasibility of the click

approach, a model reaction was carried out using the propargyl

ester derivative of 4-nitrobenzoic acid, 7. The Cu(I)-catalyzed

coupling²¹of the peracylated azido neomycin 6 with the

propargyl ester 7 (Scheme 1) was carried out in the presence

of CuSO₄and copper powder in 1:1 mixture of water and

t-BuOH at room temperature. The required triazole-containing

click product, 8, was formed cleanly and isolated in good yield

(79%). Expansions of the carbohydrate region of the ¹H NMR

spectra obtained for the per-acetylated parent neomycin, 2, the

azido derivative, 6, and the derived triazole-containing click

product, 8, are provided in Figure 2.²²The marked upfield shift

of the H_3IVresonance in the azido derivative, 6, in comparison

to the acetylated parent protected neomycin, 2, is in evidence

(Figure 2b). This shift to highfield is mirrored in the ¹³C NMR

spectrum of the compound as C_3IVresonates at 60.8 ppm in the

(22) Due to the complexity observed in spectra obtained at room

temperature,^18aall 1D and 2D ¹H NMR studies conducted on the Boc-

protected, per-acetylated neomycin B derivatives reported here have been

performed in C₅D₅N solution at 363 K (or 373 K in the case of 26). A

comparison of the ¹H NMR spectra obtained in CDCl₃and C₅D₅N solution

at room and elevated temperature is provided in Supporting Information.

(23) See for example: Szilagyi, L.; Gyorgydeak, Z. Carbohydr. Res.

1985, 143, 21-41. Compare Vignon, M. R.; Vottero, P. J. A. Tetrahedron

Lett. 1976, 17, 2445-8.

(24) Cyclenoe; NOE difference experiment, VNMR 6.1C, Varian As-

sociates.

(25) Unambiguous enhancement of the H_3IIIresonance (δ 4.73 ppm) was

also observed in this experiment (not shown).

(26) Unambiguous enhancement of the H_3Iresonance (δ 5.63 ppm) was

also observed in this experiment (not shown).

(27) The 1,4 regioselectivity of the Cu(I)-catalyzed alkyne-azide cou-

1

(20) Angyal, S. J. Chem. Ind. (London, UK) 1954, 1230-1231. Note

that a similar Fu¨rst-Plattner analysis of the other possible diastereomer

(altro epoxide) would result in azide attack at the 4-position.

(21) (a) Rostovtsev, V. V.; Green, L. G.; Fokin, V. V.; Sharpless, K. B.

Angew. Chem., Int. Ed. 2002, 41, 2596. (b) Tornøe, C. W.; Christensen,

C.; Meldal, M. J. Org. Chem. 2002, 67, 3057. (c) For a recent review of

the mechanistic and synthetic aspects of Cu(I)-catalyzed alkyne-azide

couplings, including the choice of catalysts and the regiochemical outcomes

of the reactions, see: Bock, V. D.; Hiemstra, H.; van Maarseveen, J. H.

Eur. J. Org. Chem. 2006, 51-68 and references therein.

plings in this work is verified by the ¹³C and H NMR chemical shifts of

the triazole unit in the click-coupled products obtained. We find the

resonance attributable to the triazole C_ipsoconsistently in the range δ 143-

145 ppm in the ¹³C NMR spectra of the compounds. Similarly, the triazole

CH carbon resonates in the range δ 122-127 ppm in the ¹³C NMR and the

1

triazole CH proton is observed between δ 8-8.5 ppm in the H NMR in

all cases. These values are consistent with those observed previously for

1,4-subtituted triazoles (see, for example, ref 21b) and contrast with those

reported previously for 1,5-subtituted triazoles (see, for example, Crandall,

J. K.; Crawley, L. C.; Komin, J. B. J. Org. Chem. 1975, 40, 2045-2047).

J. Org. Chem, Vol. 72, No. 6, 2007 1965

Quader et al.

1

FIGURE 2. Expansions of the carbohydrate regions of the H NMR spectra (400 MHz, 363 K, C₅D₅N) of (a) 2-hepta-O-acetate, (b) 6-hexa-O-

acetate, and (c) 8-hexa-O-acetate are shown. The changes observed in the resonances attributable to the protons of ring IV are highlighted. An

expansion of the aromatic region of the spectrum of 8-hexa-O-acetate is also provided as the inset in (c).

the external surface of the macrophage. Thus, we have chosen

to link a steroid core structure through the D-ring of 10 to

neomycin to present a mimic of the cholesterol tetracyclic

system with an unmodified 3-OH group.

FIGURE 3. Expansions of (a) the ¹H NMR spectrum (400 MHz, 363

Propargyl laurate, 9, was prepared using a standard DCC

K, C₅D₅N) of 6-hexa-O-acetate and the corresponding regions of the

coupling reaction between lauric acid and propargyl alcohol.

Copper(I)-catalyzed reaction of this ester with azidoneomycin

6 proceeded smoothly and afforded compound 13 (Scheme 1)

1D NOE spectra obtained from presaturation at (b) δ 5.13 ppm and

(c) δ 6.43 ppm, using the cyclenoe sequence²⁴with internal subtraction

(400 MHz, 363 K, C₅D₅N).

in 72% yield after purification using column chromatography.

was extended using a Wittig-Horner reaction with triethyl

Similarly, modified neomycins incorporating a steroidal lipo-

phosphonoacetate and sodium ethoxide in ethanol. Hydrolysis

philic domain were achieved using a click coupling strategy

of the resultant ethyl ester with KOH in isopropanol gave the

employing the cholesterol derivative 10 as the terminal alkyne

acid 12 (72% yield), which was then re-esterified with propargyl

synthon. The propargyl ester 10 was prepared in two steps from

alcohol, under Mitsunobu conditions, to furnish 10 in 75% yield.

the corresponding ethyl ester derivative, 11, reported previ-

The Cu(I)-mediated alkyne-azide coupling reaction of 10 with

ously.²⁸The commercially available ketone dehydroandrosterone

the azido neomycin 6 afforded the cholesterol neomycin B

hybrid 14 (Scheme 1) in 75% yield after purification.

Full assignment of the entire H NMR spectral range of the

click adducts with attached lipophilic groups is hampered by

(28) (a) Wicha, J.; Bal, K.; Piekut, S. Synth. Commun. 1977, 7, 215. (b)

Quader, S.; Boyd, S. E.; Houston, T. A.; Jenkins, I. D.; Healy, P. C. Acta

Crystallogr. 2006, E62, o162-o164.

1

1966 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

1

FIGURE 4. Expansions of the H NMR spectra (400 MHz, 363 K, C₅D₅N) of (a) 14-hepta-O-acetate and (b) the click synthon, 10. Resonances

attributable to the protons of the steroid, the ester linker, and the triazole are highlighted.

both the complexity of the aliphatic/alicyclic region and the

relative intensity of the protecting group resonances that also

occur in this region. In particular, under our experimental

conditions, the elevated probe temperature dictates the use of

traditional phase cycled 2D pulse sequences, instead of their

cleaner gradient-filtered counterparts, and hence as would be

expected, the spectral region surrounding the ¹H resonances of

the protecting groups is adversely affected by extensive T₁noise.

This inhibits full assignment of the resonances of the hydro-

carbon portions of the click adducts. However, sufficient

sensitivity and dispersion is obtained in the range above 2.5

ppm (in one- and two-dimensional experiments) to facilitate

explicit identification of the major diagnostic resonances

expected for the compounds. As in the case of the 4-nitrobenzoic

acid based click product, 8, key resonances for the triazole aryl

proton (8.25 ppm) and the methylene protons of the propargyl

linker unit (5.43 ppm) are readily identified in the high-

aminoglycoside resonances under the experimental conditions,

however. As expected, the resonances of the steroid ring protons

are largely insensitive to the changes in the linker group in the

click-coupled product. There is, however, a general shift to lower

field in the high-temperature spectrum of the fully protected

product in the C₅D₅N solvent.

Modification of the Ribose Ring. The formation of the

dehydration products observed in the reaction of neomycin B

with nucleophiles under Mitsunobu conditions renders this

method inappropriate for the synthesis of neomycin derivatives

substituted at C_5IIIunless the secondary alcohols are protected.²⁹

Selective lipophilic modification of the ribose residue of

neomycin B, without concomitant modification of the ido ring,

could be accomplished, however, via a procedure based in part

on the previously reported selective tripsylation of the C_5III

primary hydroxyl with 2,4,6-triisopropylbenzenesulfonyl chlo-

ride (TPSCl) (Scheme 3).³⁰N-Boc-protected neomycin B 2 (1

equiv) was treated with excess TPSCl (28 equiv) in pyridine to

obtain the tripsylated derivative 15 (61%). Treatment of 15 with

sodium azide in DMF at 100 °C (18 h) afforded the ribo-

substituted azido neomycin B, 16, in quantitative yield. Cu-

catalyzed alkyne-azide coupling of the archetypal click synthons

7, 9, and 10 to 16 afforded the corresponding triazole derivatives

17 (75%), 18 (71%), and 19 (69%) in good yield.

1

temperature H NMR spectrum of the lauric acid adduct, 13-

hexa-O-acetate. Similarly, downfield shifts are observed for the

resonances attributable to H_3IVand H_4IV(δ 5.38 and 5.71 ppm),

and these resonances correlate with resonances at 61.3 and 69.8

ppm, respectively, in the ¹³C NMR spectrum of the same

material (HMQC). Figure 4 highlights the diagnostic resonances

attributable to the cholesterol-derived portion of the per-

acetylated click product, 14-hepta-O-acetate. The doublet of

doublets assigned to H₂₀of the steroid side-chain, the broad

singlet of the OCH₂steroid/neomycin linker group, the AB

systems observed for the H₄and H₁₆steroid methylene pairs,

and the singlet resonance of the aromatic proton of the triazole

unit are all clearly evidenced. The resonances attributable to

H₆and H₃of the acetylated steroid are co-incident with

1

The diagnostic features of the high-temperature H NMR

spectra of the ribo-substituted azido neomycin B, 16-hexa-O-

(29) (a) Hanessian, S.; Tremblay, M.; Swayze, E. E. Tetrahedron 2003,

59, 983. (b) Liang, F.-S.; Wang, S.-K.; Nakatani, T.; Wong, C.-H. Angew.

Chem., Int. Ed. 2004, 43, 6496-6500.

(30) Michael, K.; Wang, H.; Tor, Y. Bioorg. Med. Chem. 1999, 7, 1361-

1371.

J. Org. Chem, Vol. 72, No. 6, 2007 1967

Quader et al.

1

FIGURE 5. Expansions of the H NMR spectra (400 MHz, 363 K, C₅D₅N) of (a) 16-hexa-O-acetate and (b) 17-hexa-O-acetate. The changes

observed in the resonances attributable to the protons of ring III are highlighted. An expansion of the aromatic region of the spectrum of 17-hexa-

O-acetate is also provided as the inset in (b).

SCHEME 3^a

^aReagents and conditions: (a) TPSCI (28 equiv), pyridine, rt, 18 h; (b) NaN₃, 100 °C, DMF, 8 h; (c) R′COO-CH₂C′′ CH (2 equiv), t-BuOH/H₂O (1:1),

Cu/CuSO₄, rt, 18 h.

acetate, and the derived click product 17-hexa-O-acetate (Scheme

3) are highlighted in Figure 5. It can be seen that the resonances

supra). The ¹³C NMR resonance attributable to C_5IIIalso exhibits

a dramatic upfield shift in the spectrum of the azido derivative

16-hexa-O-acetate (δ 51.94 ppm, cf. C_5III63.9 ppm in 2-hepta-

O-acetate). This shift is again consistent with the chemical shift

of the azide bearing carbon in compound 6-hexa-O-acetate and

with literature values.

On coupling with the click synthon 7, the initially high field

resonances of H_5IIIand H_5′IIIshow a marked downfield shift in

17-hexa-O-acetate (δ 5.15 and 4.94, Figure 5b), resonating 0.2-

assignable to protons H_5IIIand H

in the azido derivative

5′III

16-hexa-O-acetate (δ 3.97 and 3.76 ppm, Figure 5a) become

increasingly dispersed and resonate 0.7-1 ppm upfield of the

corresponding resonances in the per-acetylated parent, 2-hepta-

O-acetate. These shifts are consistent with the ∆δ values

observed for the H_3IVring proton of azido compound 6-hexa-

O-acetate, when compared with the parent compound (vide

1968 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

FIGURE 6. Expansions of the ¹H NMR spectra (400 MHz, 363 K, C₅D₅N) of (a) 3-penta-O-acetate and (b) 4-tetra-O-acetate. The changes observed

in the resonances attributable to the protons of ring II are highlighted.

0.5 ppm to lower field of the antecedent resonances of the per-

acetylated parent compound. The resonances in the lauric acid

derived analogue, 18-hexa-O-acetate (δ 5.13 and 4.93 ppm),

and the cholesterol adduct, 19-hepta-O-acetate (δ 5.11 and 4.98),

similarly show distinctive downfield shifts, indicative of triazole

formation adjacent to C_5III. However, as was observed in the

case of the ido substituted analogues, the ¹³C NMR shift of C_5III

is less sensitive to the conversion of the azide function to a

triazole derivative, C_5IIIresonating at δ51.9 and 52.4 ppm in

compounds 16-hexa-O-acetate and 19-hepta-O-acetate, respec-

tively.

1

In addition to the notable changes in the H NMR chemical

shifts of the H_5IIIand H

protons, a consistent diagnostic

5′III

feature of adducts isolated from click coupling reactions

conducted on the ribo substituted azido neomycin B, 16, is the

appearance of the resonances attributable to the OCH₂methylene

protons of the propargyl alcohol derived unit of the click

synthon. In the spectra of the per-acetylated derivatives such

as 17-hexa-O-acetate (Figure 5b), the resonances of these

diastereotopic protons diverge in chemical shift and appear as

distinct AB systems (400 MHz). This contrasts with the adducts

formed from ido substituted azido neomycin B, 6 (Figure 2c),

where in all cases the methylene protons of the per-acetylated

systems show co-incident resonant frequencies.

FIGURE 7. Expansions of the aziridine region of (a) the HMQC

spectrum and (b) the HMBC spectrum obtained for 4-tetra-O-acetate

(400 MHz, C₅D₅N, 363 K). The corresponding region of ¹H NMR

spectrum is provided as an axis reference.

6. It is noteworthy that an NH resonance, present in the mono-

anhydro species (Figure 6a), is absent in the spectrum of the

bis-anhydro derivative, 4-tetra-O-acetate (Figure 6b). In this

instance, the second dehydration step clearly proceeds with one

of the protected amine nitrogens serving as the intramolecular

nucleophile, rather than a second hydroxylic oxygen nucleophile.

Modification of the Steptamine Ring. As discussed above,

a second major dehydration product, 4, was isolated from

reaction mixtures containing the protected parent species, 2, and

the Mitsunobu reagents TPP and DIAD (Scheme 1). Mass

spectral analysis of the per-acetylated derivative confirmed a

tetra-O-acetate, supporting the formation of a stable bis-anhydro

1

The marked changes observed in the H NMR and ¹³C NMR

derivative of the parent aminoglycoside. Subsequent ¹H and ¹³

C

resonances of the streptamine residue (ring II) locates the

resultant aziridine within this ring. Distinct shifts are observed

for the resonances of the H_5IIand the H_1IIand H_6IIprotons in

particular. H_5IIresonates 0.5 ppm downfield from its antecedent,

while H_1IIand H_6IImove 1.2 and 2.1 ppm upfield, respectively.

Further evidence of aziridine ring formation at C_6IIand C_1IIis

obtained from the ¹³C NMR chemical shifts of the C_6IIand C_1II

carbons. Unambiguous cross-peaks are observed in the HMQC

NMR analysis of the isolated, per-acetylated aminoglycoside,

4-tetra-O-acetate, revealed the formation of an aziridine ring in

ring II, with a Boc-protected nitrogen bridging C_6IIand C_1II, in

addition to the previously observed epoxide at C_3IVand C_4IVof

the ido ring.

Expansions of the ¹H NMR spectrum of 4-tetra-O-acetate and

its mono-anhydro precursor 3 (Scheme 1) are provided in Figure

J. Org. Chem, Vol. 72, No. 6, 2007 1969

Quader et al.

FIGURE 8. (a) NOE enhancements observed in cyclenoe experiments conducted on 4-tetra-O-acetate and summarizing intraresidue enhancements

3

observed in experiments focused on ring IV. (b) Preferred conformation of ring II. J_HHcouplings, obtained in the simulation, are provided in the

table inset. R_IO and R_IIIO represent glycosidic linkages to rings I and III.

spectrum of 4-tetra-O-acetate linking H_1IIand H_6IIwith ¹³C

resonances at δ 38.1 and 38.5 ppm, respectively (Figure 7a).

These shifts are in accord with those observed previously in

Boc-protected aziridines.³¹Furthermore, the HMBC spectrum

(Figure 7b) of 4-tetra-O-acetate shows distinct cross-peaks

correlating H_1IIand H_6IIwith a single carbamate CdO function

resonating at 162 ppm. This CdO resonates some 5 ppm

downfield of all other carbamate (carbonyl) resonances observed

both in this species and other compounds studied in this work

and is consistent with literature values recorded for the car-

bamate functions of Boc-protected aziridines.³¹The two HMBC

cross-peaks linking both H_1IIand H_6IIwith this carbamate

resonance confirms the formation of the three-membered

aziridine ring at this position.

phosphorane intermediate is formed. This ring may stabilize

the all-axial conformation of the deoxystreptamine required for

aziridine ring formation and/or facilitate transfer of the tri-

phenylphosphinoxy leaving group from the primary hydroxyl

of ribose to the secondary hydroxyl of the deoxystreptamine

(Scheme 4). From a study of molecular models, the arrangement

of the two oxygen atoms about the phosphorus could be either

diaxial or axial-equatorial, the latter being less crowded (these

two forms can equilibrate via Berry pseudorotation³⁴). Phos-

phoranes of this type are well-known intermediates in the

Mitsunobu reaction.³⁵The corresponding 14-membered ring

phosphorane joining the 5-position of ribose to the 3-position

of the gluco ring is highly strained and much less likely to form.

Attempts to improve the yield of 4, by increasing the

proportion of TPP and DIAD added to 2 from 4 to 6 equiv,

resulted in essentially no change in the isolated yield of 4 (48%,

6:6:1 TPP/DIAD/2; cf. 45%, 4:4:1 TPP/DIAD/2). Mass spectral

analysis of the product mixture obtained in the reaction revealed

the presence of significant amounts (18%) of a compound with

a molecular ion corresponding to a DIAD-derived hydrazine

adduct (LRESMS [M + Na⁺] ) 1387) of the initially formed

bis-anhydro derivative 4. In order to suppress the formation of

DIAD derived hydrazine adducts and thus improve the yield of

4, the bulkier di-tert-butyl azodicarboxylate (DBAD) was

substituted for DIAD in the synthesis of 4. Thus, 2 was treated

with 6 equiv of TPP and DBAD in toluene at room temperature

(12 h). This procedure resulted in a 10% improvement in the

isolated yield of 4 (55%) and only traces of putative DBAD-

derived hydrazine adducts were observed in these reactions.

1D NOE experiments conducted on 4-tetra-O-acetate (Scheme

1) support an average ^OH₁conformation of ring IV, as observed

in the mono-anhydro precursor, 3-penta-O-acetate. Enhancement

of the H_5IVresonance results from selective saturation of either

H_4IVor H_1IV. Saturation of H_3IVresults in enhancement of H_2IV

as expected (Figure 8a). NOE experiments conducted on ring

II, however, yielded ambiguous results due to the near co-

incident overlap of the H_3IIand H_4IIresonances.³²Simulation

of the ¹H NMR resonances of the ring II spin system, for 4-tetra-

3

O-acetate and related compounds (Scheme 5), yielded J_HH

vicinal coupling constants similar to those observed by Crotti

and co-workers in their studies³³on aziridine-containing pyrans.

In particular, we typically observe values of ∼0.1 and 5.0 Hz

3

for the J_1II,2aIIand J_1II,2bIIcoupling constants of compounds

synthesized that are isostructural in ring II. These distinct values

suggest that ring II averages to a ⁴H₃conformation (Figure 8b),

as the alternate ³H₄conformation would be expected to exhibit

similar values for the ³J_1II,2aIIand ³J_1II,2bIIcouplings.³³Further-

more, the values fitted for the other ³J couplings of ring II, for

this compound and related aziridine-containing compounds

synthesized in this work, are also consistent with those expected

The bis-anhydro neomycin B derivative 4 invites further

synthetic modification. The ring opening of the epoxide and/or

aziridine functions permits modification of two sugar residues

in the neomycin B framework. Modification of ring II, a key

pathogen/target binding residue,³⁶in particular offers consider-

able potential for significantly affecting the resultant aminogly-

coside’s antibiotic activity. Accordingly, we have undertaken

preliminary studies exploring the ring opening of the epoxide

and/or the aziridine rings of compound 4.

4

for the H₃conformation.

An interesting observation is that the formation of the

aziridine ring takes place on the deoxystreptamine ring but not

on the gluco ring. Also, when the 5-position of the (remote)

ribose is blocked (as the 4-nitrobenzoate ester), aziridine ring

formation does not occur. This suggests that the 5-position of

ribose takes part in the mechanism of formation of the aziridine

ring. A possible explanation is that a cyclic 10-membered ring

HPLCMS (MeCN/H₂O 75:25) analysis of a per-acetylated

product mixture obtained after treatment of the bis-anhydro

(34) (a) Berry, R. S. J. Chem. Phys. 1960, 32, 933. (b) Gallagher, M. J.;

Jenkins, I. D. In Topics in Stereochemistry; Eliel, E. L., Allinger, N. L.,

Eds.; New York: Wiley-Interscience, 1968; Vol. 3, p 64.

(31) See, for example: Gardiner, J. M.; Loyns, C. R. Tetrahedron 1995,

51, 11515-11530.

(35) (a) von Itzstein, M.; Jenkins, I. D. Aust. J. Chem. 1983, 36, 557-

563. (b) von Itzstein, M.; Jenkins, I. D. J. Chem. Soc., Perkin Trans. 1

1986, 437-445. (c) von Itzstein, M.; Jenkins, I. D. J. Chem. Soc., Perkin

Trans. 1 1987, 2057-2060. (d) Camp, D.; Jenkins, I. D. J. Org. Chem.

1989, 54, 3045-3049; 3049-3054.

(32) Sample spectra obtained in the NOE experiments conducted on ring

IV are provided in Supporting Information.

(33) Crotti, P.; Di Bussolo, V.; Favero, L.; Pineschi, M. Tetrahedron

1997, 53, 1417-1438.

(36) Magnet, S.; Blanchard, J. S. Chem. ReV. 2005, 105, 477-497.

1970 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

SCHEME 4

SCHEME 5^a

^aReagents and conditions: (a) (i) 1 equiv NaN₃, 50 °C, DMF, 2 d, (ii) Ac₂O, C₅H₅N, DMAP; (b) (i) excess NaN₃, 50 °C, DMF, 1 d, (ii) Ac₂O, C₅H₅N,

DMAP; (c) O₂NC₆H₄COO-CH₂C′′CH (2 equiv), t-BuOH/H₂O (1:1), Cu/CuSO₄, rt, 18 h; (d) excess NaN₃, 50 °C, DMF, 1 d.

neomycin derivative 4 with 1 equiv of sodium azide in DMF

(Scheme 5) revealed a product mixture comprising 4-tetra-O-

acetate (32%), a diazido adduct, 20-penta-O-acetate (11%), and

two possible mono-azido adducts, 21-penta-O-acetate and 22-

tetra-O-acetate (40% and 15%, respectively). Subsequent at-

tempts at product purification showed that although the diazido

adduct 20 and the mono-azido adduct 21 could be isolated, the

second mono-azido adduct 22 coeluted with the residual starting

material in all solvent systems examined. Attempts to separate

the species after per-acetylation of the crude reaction mixture

met with only limited success. The diazido adduct 20-penta-

O-acetate was again successfully isolated; however, analytical

quantities of the mono-azido adducts were obtained only after

preparative HPLC. The mono-azido species resulting from ring

J. Org. Chem, Vol. 72, No. 6, 2007 1971

Quader et al.

1

FIGURE 9. Expansions of the carbohydrate regions of the H NMR spectrum (400 MHz, C₅D₅N, 363 K) of (a) 20-penta-O-acetate and (b)

24-tetra-O-acetate. NOE enhancements resulting from selective saturation of H_2IIeqare provided for each compound in (c) and (d). NOE enhancements

resulting from selective saturation of H_2IIaxare provided for each compound in (e) and (f).

opening of the ring II aziridine, 22-tetra-O-acetate, has not been

obtained in sufficient quantity for full characterization (¹H NMR

only). However, a separable mixture of the bis-anhydro derivate,

4-tetra-O-acetate, and the click adducts 23-penta-O-acetate and

24-tetra-O-acetate was obtained when a partially purified

mixture of 4-tetra-O-acetate, 21-penta-O-acetate, and the uniso-

lated 22-tetra-O-acetate was treated with the click synthon 7

under our standard conditions. The click coupling proceeds

quantitatively with respect to the initial concentration of azido

species present in the reaction mixture in both cases. No

degradation of the aziridine-containing ring II was observed in

the formation of 23-penta-O-acetate, and no degradation of the

epoxide-containing ring IV was observed in the formation of

24-tetra-O-acetate under the conditions of the click coupling

reaction.

the shift of the H_1IIresonance 1.35 ppm downfield with respect

to the aziridine precursor. Furthermore, C_1IIresonates at δ 57.3

ppm in the ¹³C NMR (cf. 50.5 ppm in 2-hepta-O-acetate). The

migration of the Boc protected amine to C_6IIis confirmed by

the cross-peak correlating H_6IIand the NH proton in the COSY

spectrum.³⁸The attack by azide at C_1II, rather than C_6II, is not

surprising. C_1IIis the least hindered site for attack, and moreover,

being adjacent to the methylene group, this site would be favored

on electronic grounds as the transition state for attack by azide

would result in the development of a small positive charge at

the aziridine carbon. Attack would therefore be favored by an

adjacent CH₂rather than a CHOR group. It was surprising,

however, that the product of azide attack did not retain the ⁴C₁

conformation expected from the opening of the ⁴H₃conformer.

The initial trans-diaxial arrangement of the C_1IIazido and C_6II

NHBoc groups is unobserved in the product. Instead, ring II of

The ring opening of the epoxide function of ring IV by azide

ion in both the mono-azido adduct 21 and the diazido adduct

20 (Scheme 5) proceeded in a manner directly analogous to

that observed in the reaction of the mono-anhydro neomycin

derivative 4, as expected. The talo epoxide of ring IV of 4

undergoes regioselective ring opening by azide ion at C_3IVin

1

20-penta-O-acetate averages to a C₄conformation, in which

the C_1IIazido and C_6IINHBoc groups are diequatorial, while

1

the substituents on C_3II, C_4IIand C_5IIare all axial. The C₄

conformation of ring II of 20-penta-O-acetate is evidenced in

the ¹H NMR spectrum. The H_1IIresonance of 20-penta-O-acetate

is well-separated from surrounding resonances in the C₅D₅N

solvent at 363 K and shows couplings of 4, 10, and 10 Hz to

neighboring protons, suggesting an axial disposition of the

proton. Furthermore, selective irradiation of the H_2IIax(δ 2.12

ppm) and H_2IIeq(δ 2.36 ppm) resonances of 20-penta-O-acetate

engenders NOE at H_3IIin both cases (Figure 9). Significant

enhancement of the H_1IIand NH_(3II)resonances is only observed

in the case of selective irradiation of the H_2IIeqresonance;

however, further supporting the mutually trans-diaxial disposi-

tions of H_1II, H_2IIax, and the NH_(3II)Boc group. It is also

noteworthy that the resonances of H_1IIIand H_2IIIshow marked

shifts in 20-penta-O-acetate relative to their antecedents in the

1

both cases. The H and ¹³C NMR resonances of the ring IV

fragments of the per-acetylated derivatives of 20 and 22 are in

close agreement with those observed for 6-hexa-O-acetate.

Similarly, chemical shift, coupling constant, and NOE experi-

ments focused on the aziridine containing ring II of the 21-

and 23-penta-O-acetates showed results in accord with those

observed for the bis-anhydro parent species 4-tetra-O-acetate.

4

The H₃conformations are observed for ring II in both cases.

Analysis of the ¹H and ¹³C NMR spectra obtained for diazido

adduct 20-penta-O-acetate³⁷revealed an interesting outcome in

the ring opening of the aziridine function of ring II by azide

ion. Azide substitution occurs at C_1II, resulting in a net migration

of the amine function from C_1IIin the parent neomycin to C_6II

in these derivatives. Azide substitution at C_1IIis supported by

(38) (a) Separate crosspeaks for the H_6II-NH_(6II)and H_3II-NH_(3II)

correlations are clearly evident in the 2D COSY experiment (400 MHz,

363 K, C₅D₅N); see Supporting Information. (b) Migratory, aziridine

formation and ring opening has been observed in streptamine systems

before: Ikeda, D.; Horiuchi, Y.; Kondo, S.; Umezawa, H. J. Antibiot. 1980,

33, 1281-1288.

(37) The diazido adduct 20, obtained in low yield in the reaction of 4

with equimolar sodium azide, was obtained quantitatively on treatment of

4 with excess sodium azide.

1972 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

parent neomycin, although no direct chemical modification has

taken place in this ring. A similar picture emerges from NMR

spectroscopic studies conducted on derivatives that are iso-

structural at ring II, i.e., 22-tetra-O-acetate and 25-penta-O-

acetate, as well as the triazole derivative 24-tetra-O-acetate

(Figure 9). The body of data suggests a significant conforma-

tional change accompanies the migration of the amine function

of derivatives of neomycin B. Indeed, by employing a strategy

based on the bis-anhydro derivative 4, highly modified deriva-

tives of the neomycin B parent framework can be obtained. For

example, treatment of the isolated click product 23-penta-O-

acetate (Scheme 5) with excess sodium azide under the standard

conditions yields an aminoglycoside derivative, 25-penta-O-

acetate, bearing a hydrolyzable lipophilic function at C_3IVand

a transformable azido functionality located in ring II. Moreover,

ring II has an orientation of functional groups (three axial and

two equatorial) different from that of the parent deoxy-

streptamine ring (all groups equatorial) of neomycin. Further-

more, the anhydro neomycin 3 can now be produced efficiently

and in high yield and provides an excellent building block for

the preparation of a library of neomycin analogues bearing a

range of functionality at the 3-position of the ido moiety. The

biological activity of the modified neomycins reported here is

currently being investigated.

1

in ring II, with this ring averaging to a C₄conformation, in

4

contrast to the C₁conformation of the neomycin parent, even

though this conformation places three bulky substituents in axial

1

positions. We suggest that the C₄conformation is stabilized

by a H-bonding interaction between the axial NHBoc group on

C_3IIand the axial oxygen on C_5II. Recent theoretical calcula-

tions³⁹and a literature example⁴⁰suggest that H-bonding

interactions can affect conformational preferences in these

systems.

Experimental Section

1,3,2′,6′,2′′′,6′′′-Hexa-N-(tert-butoxycarbonyl)-neomycin B (2).

Triethylamine (TEA) (7 mL) and methanol (10 mL) were added to

a stirred solution of neomycin B sulfate 1 (2 g, 2.2 mmol) in water

(10 mL). Di-tert-butyl dicarbonate (5 g, 22 mmol) was then added,

and the resultant reaction mixture was stirred at elevated temperature

(55 °C, 16 h). Methanol was removed by evaporation and the

residue was partitioned between ethyl acetate (200 mL) and water

(100 mL). The aqueous layer was extracted with fresh ethyl acetate

(2 × 50 mL) and the combined organic layers were dried (Na₂-

SO₄) and concentrated in vacuo. Purification by flash chromatog-

raphy on silica (DCM/acetone 3:2) afforded 2 as an amorphous

white solid (2.42 g, 90%). LRMS (ESI): calculated for (M + Na⁺)

C₅₃H₉₄N₆O₂₅1237.61, found 1237.6.

Attempts were made to improve the regioselectivity of the

initial azide addition to 4. However, essentially no improvement

in yield or selectivity was observed when either catalytic

quantities of cerium chloride were added to the reaction mixture

or when the reactions were carried out using trimethylsilylazide

(TMSA)⁴¹in place of NaN₃.⁴²Treatment of 4 with neat

propylamine at room temperature for 16 h did, however, afford

a 47% yield of the product obtained from selective ring opening

of the aziridine function of ring II, 26. The starting material, 4,

was recovered in 50% yield after purification. Only traces of

the product resulting from ring opening of both the aziridine

and epoxide functions were detected in the product mix. The

alternate epoxide opened species was undetected in the mixture.

Investigations into alternate methods for improving the yield

and regioselectivity of the ring opening reactions are ongoing

in our laboratory.

General Procedure for the Per-acetylation. 2-Hepta-O-

acetate.^18aCompound 2 (25 mg, 0.02 mmol) was treated with acetic

anhydride and pyridine (1:1, v/v, 2 mL) in the presence of a catalytic

amount of N,N-dimethylaminopyridine (DMAP) at room temper-

ature (4 h). The solvent was removed in vacuo and the residue

was purified by flash chromatography (DCM/MeOH 98:2). This

afforded 2-hepta-O-acetate as a white amorphous solid (30 mg,

1

98%). H NMR (400 MHz, pyridine-d₅, 363 K): δ 6.97 (1H, d,

J ) 8 Hz, NH_II), 6.79 (1H, d, J ) 8 Hz, NH_II), 6.70 (1H, br dd,

NH_IV), 6.57 (1H, br dd, NH_I), 6.48 (1H, d, J ) 10 Hz, NH_I), 6.09

(1H, d, J ) 9.5 Hz, NH_IV), 6.00 (1H, br s, H_1I), 5.64 (1H, dd, J )

9.5, 10.8 Hz, H_3I), 5.60 (1H, d, J ) 2.5 Hz, H_{1 III}), 5.52 (1H, dd,

J ) 3, 3 Hz, H_3IV), 5.41 (1H, dd, J ) 9.5, 10 Hz, H_4I), 5.41 (1H,

dd, J ) 2.5, 5 Hz, H_{2 III}), 5.24 (1H, ddd, J ) 3, 2, 1 Hz, H_4IV),

5.23 (1H, dd, J ) 10.3, 9.3 Hz, H_6II), 5.19 (1H, d, J ) 2 Hz, H_1IV),

4.75 (2H, m, H_{3 III}, H_{5 III}), 4.66 (1H, dd, J ) 11.9, 5.3 Hz, H_5III’),

4.61 (1H, ddd, J ) 4, 4, 10 Hz, H_5I), 4.51-4.43 (3H, br m, H_2I,

H_5IV, H_4III), 4.30 (1H, br m, H_2IV), 4.14-3.9 (4H, br m, H_1II, H_3II,

H_4II, H_5II), 3.82 (1H, ddd, J ) 7, 7, 14 Hz, H_6IV), 3.72 (2H, m, H_6I,

H_6I’), 3.61 (1H, ddd, J ) 5, 7, 14 Hz, H_6IV’), 2.41 (3H, s, -OCOCH₃),

2.40 (1H, m, H_2IIeq), 2.35 (3H, s, -OCOCH₃), 2.29 (3H, s,

-OCOCH₃), 2.15 (3H, s, -OCOCH₃), 2.14 (3H, s, -OCOCH₃), 2.10

(3H, s, -OCOCH₃), 2.04 (3H, s, -OCOCH₃), 2.0 (1H, m, H_2IIax),

1.68 (9H, s, -COC(CH₃)₃), 1.63 (9H, s, -COC(CH₃)₃), 1.58 (9H, s,

-COC(CH₃)₃), 1.57 (18H, s, 2 × -COC(CH₃)₃), 1.56 (9H, s, -COC-

(CH₃)₃). ¹³C {¹H} NMR (100 MHz, pyridine-d₅, 363 K): δ 171.5-

169.3⁴³(7 × OAc CdO), 157.5-156.7⁴³(6 × Boc CdO), 108.0

(C_1III), 99.2 (C_1IV), 98.3 (C_1I), 83.5 (C_5II/C_4II), 79.9 (C_4II/C_5II), 79.9

(C_4III), 79.6-78.8⁴³(6 × Boc C_q), 76.9 (C_6II), 76.5 (C_3III), 75.2

(C_2III), 73.7 (C_5IV), 73.0 (C_3I), 70.9 (C_4I), 70.0 (C_3IV), 69.8 (C_5I),

67.5 (C_4IV), 63.9 (C_5III), 54.3 (C_2I), 50.5 (C_2IV, C_1II, C_3II), 41.9 (C_6I),

41.4 (C_6IV), 35.4 (C_2II), 28.9-27.6⁴³(6 × Boc (CH₃)₃), 21.8-20.7⁴³

Conclusion

We have demonstrated the selective modification of the ido-

(IV), ribo- (III), and streptamine (II) rings of the neomycin B

framework utilizing two different yet complementary synthetic

approaches. Utilizing these strategies we have prepared a number

(39) de Oliveira, P. R. and Rittner, R. Spectrochim. Acta., Part A 2005,

62, 30-37.

(40) Sundaralingam, M.; Brennan, R.; Swaminathan, P.; Haromy, T. P.;

Drendel, W. B.; BeMiller, J. N.; Chmielewski, M.; Cerretti, D. P.

J. Carbohydr. Chem. 1982, 1, 85-103.

(41) Sabitha, G.; Babu, R. S.; Rajkumar, M.; Yadav, J. S. Org. Lett.

2002, 4, 343-345.

(42) (a) Wu, J.; Hou, X. J. Org. Chem. 1999, 1344-1348. (b)

Chandrasekhar, M.; Sekar, G.; Singh, V. K. Tetrahedron Lett. 2000, 41,

10079-10083.

(43) Recorded at room temperature.

J. Org. Chem, Vol. 72, No. 6, 2007 1973

Quader et al.

(7 × OAc CH₃). LRMS (ESI): calculated for (M + Na⁺)

29.0⁴³(6 × Boc (CH₃)₃), 22.0-21.3⁴³(5 × OAc CH₃). HRMS

(ESI): calculated for (M + H⁺) C₆₃H₁₀₂N₆O₂₉1407.67695, found

1407.67615.

C₆₇H₁₀₈N₆O₃₂1531.6, found 1531.6.

3′′′,4′′′-Anhydro-1,3,2′,6′,2′′′,6′′′-hexa-N-(tert-butoxycarbonyl)-

neomycin B (3)^18aand 3′′′,4′′′-Anhydro-3,2′,6′,2′′′,6′′′-penta-N-

(tert-butoxycarbonyl)-1,6-[N-(tert-butoxycarbonyl)epimino]-1-

deamino-6-deoxyneomycin B (4). DIAD (330 µL, 1.6 mmol) was

added slowly to an ice-cold solution of 2 (500 mg, 0.41 mmol)

and TPP (430 mg, 1.6 mmol) in dry toluene (5 mL) under a dry N₂

atmosphere. The mixture was stirred at room temperature (18 h)

and then evaporated to dryness. Column chromatography on silica

(DCM/MeOH 97:3 f 95:5) afforded 4 (220 mg, 45%) [HRMS

(ESI) calculated for (M + H⁺) C₅₃H₉₀N₆O₂₃1179.61356, found

1179.60897] and 3 (160 mg, 33%) [HRMS (ESI) calculated for

(M + Na⁺) C₅₃H₉₂N₆O₂₄1219.60607, found 1219.60032] as a white

amorphous solid.

Optimal Conditions for 3′′′,4′′′-Anhydro-1,3,2′,6′,2′′′,6′′′-hexa-

N-(tert-butoxycarbonyl)-neomycin B (3). DIAD (170 µL, 0.82

mmol) was added slowly to a stirred ice-cold solution of 2 (500

mg, 0.41 mmol) and TPP (215 mg, 0.82 mmol) in dry toluene (3

mL) and the resultant reaction mixture was stirred at room

temperature under a dry N₂atmosphere (12 h). A second aliquot

of a toluene solution of TPP (215 mg, 0.82 mmol, 2 mL) was then

added, followed by the addition of a second portion of DIAD (170

µL, 0.82 mmol) and the reaction was stirred for a further 12 h prior

to removal of the solvent in vacuo. Column chromatography of

the residue on silica (DCM/MeOH 97:3 f 95:5) afforded 3 as an

amorphous white solid (305 mg, 62%).

Optimal Condition for 3′′′,4′′′-Anhydro-3,2′,6′,2′′′,6′′′-penta-

N-(tert-butoxycarbonyl)-1,6-[N-(tert-butoxycarbonyl)epimino]-

1-deamino-6-deoxyneomycin B (4). DBAD (227 mg, 0.99 mmol)

was added slowly, under a dry N₂atmosphere, to a stirred ice-cold

solution of 2 (200 mg, 0.16 mmol) and TPP (258 mg, 0.99 mmol)

in dry toluene (3 mL). The reaction mixture was stirred overnight

at room temperature and then concentrated in vacuo. Column

chromatography of the resultant residue on silica (DCM/MeOH 97:

3 f 95:5) afforded compound 4 as an amorphous white solid (107

mg, 55%).

3′′′,4′′′-Anhydro-1,3,2′,6′,2′′′,6′′′-hexa-N-(tert-butoxycarbonyl)-

5′′-deoxy-5′′-(4-nitrobenzoyl)-neomycin B (5). DIAD (330 µL,

1.65 mmol) was added slowly to a cold solution of 2 (200 mg,

0.165 mmol), TPP (430 mg, 1.65 mmol), and 4-nitrobenzoic acid

(330 mg, 2.0 mmol) in dry solvent (2 mL toluene + 1 mL THF)

under a dry N₂atmosphere. The mixture was stirred at room

temperature (18 h) and then evaporated to dryness. Column

chromatography on silica (DCM/MeOH 97:3 f 95:5) afforded 5

(187 mg, 85%) as a white amorphous solids. HRMS (ESI):

calculated for (M + Na⁺) C₆₀H₉₅N₇O₂₇1368.61736, found

1368.61214.

4-Tetra-O-acetate. Per-acetylation of compound 4 was carried

out exactly as described above for 2-hepta-O-acetate, affording

4-tetra-O-acetate as an amorphous white solid in quantitative yield.

¹H NMR (400 MHz, pyridine-d₅, 363 K): δ 6.79 (1H, br dd, NH_IV),

6.60 (1H, br dd, NH_I), 6.55 (1H, d, J ) 9 Hz, NH_I), 6.26 (1H, d,

J ) 8.5 Hz, NH_II), 6.18 (1H, d, J ) 9.5 Hz, NH_IV), 6.01 (1H, br s,

H_1III), 5.69 (1H, d, J ) 5.2 Hz, H_2III), 5.64 (1H, d, J ) 3.8 Hz,

H_1I), 5.54 (1H, dd, J ) 9.4, 10.8 Hz, H_3I), 5.37 (1H, dd, J ) 10,

9.5 Hz, H_4I), 4.84 (1H, d, J ) 3.3 Hz, H_1IV), 4.80 (1H, dd, J ) 5.3,

7.1 Hz, H_3III), 4.64 (2H, m, H_5III, H_5′III), 4.54 (1H, ddd, J ) 4.6,

4.6, 7 Hz, H_4III), 4.50 (1H, m, H_5II), 4.48-4.37 (3H, m, H_2IVH_2I,H_5I),

4.33 (1H, dd, J ) 6.3,6.3 Hz, H_5IV), 4.12 (2H, m, H _3II,H_4II), 3.76-

3.67 (4H, m, H_6IV, H_6IV′, H_6I, H_6I’), 3.50 (1H, dd, J ) 5.3, 3.8 Hz,

H_3IV), 3.34 (1H, d, J ) 3.8 Hz, H_4IV), 3.12 (1H, dd, J ) 6.4, 4.7

H_6II), 2.80 (1H, dd, J ) 5.1, 6.4 Hz, H_1II), 2.42 (3H, s, -OCOCH₃),

2.30 (1H, m, H_2IIb), 2.28 (3H, s, -OCOCH₃), 2.15 (3H, s,

-OCOCH₃), 2.06 (3H, s, -OCOCH₃), 2.05 (1H, m, H_2IIa), 1.65 (9H,

s, -COC(CH₃)₃), 1.61 (9H, s, -COC(CH₃)₃), 1.60 (9H, s, -COC-

(CH₃)₃), 1.58 (18H, s, 2 × -COC(CH₃)₃), 1.53 (9H, s, -COC(CH₃)₃).

¹³C {¹H} NMR (100 MHz, pyridine-d₅, 363 K): δ 171.4-170.0⁴³

(4 × OAc CdO), 162.4 (Boc CdO), 159.5-154.4 (5 × Boc Cd

O), 105.0 (C_1III), 99.7 (C_1IV), 99.0 (C_1I), 81.6 (C_4III), 80.6-78.0⁴³

(6 × Boc C_q), 77.0 (C_3III), 76.6 (C_4II), 76.4 (C_2III), 74.1 (C_5II), 73.0

(C_3I,C_5IV), 71.4 (C_4I), 70.8 (C_5I), 65.0 (C_5III), 54.9 (C_2I), 52.8 (C_4IV),

52.5 (C_3IV), 49.4 (C_3II), 46.6 (C_2IV), 43.7 (C_6IV), 42.6 (C_6I), 38.5

(C_6II), 38.1 (C_1II), 28.4 (C_2II), 29.3-28.0 (6 × Boc (CH₃)₃), 20.8-

20.6 (4 × OAc CH₃). HRMS (ESI): calculated for (M + H⁺)

C₆₁H₉₈N₆O₂₇1347.65581, found 1347.65496.

5-Tetra-O-acetate. Per-acetylation of compound 5, according

to the general procedure, provided 5-tetra-O-acetate. ¹H NMR (400

MHz, pyridine-d₅, 363 K): δ 8.57 (2H, m, ArH _AA′), 8.49 (2H, m,

ArH _XX′), 7.03 (1H, d, J ) 8.4 Hz, NH_II), 6.84 (2H, br dd, NH_IV),

6.72 (1H, d, J ) 7.9 Hz, NH_II), 6.52 (1H, d, J ) 9.8 Hz, NH_I),

6.36 (2H, br dd, NH_I), 6.16 (1H, d, J ) 9.6 Hz, NH_IV), 6.04 (1H,

br s, H_1I), 5.61 (1H, d, J ) 1.9 Hz, H_1III), 5.59 (1H, dd, J ) 9.5,

10.6 Hz, H_3I), 5.47 (1H, dd, J ) 1.6, 5.1 Hz, H_2III), 5.20 (2H, m,

H_6II,H_4I), 4.71 (1H, dd, J ) 3.0, 12.0 Hz,H_5III), 4.80-4.78(2H, m,

3-Penta-O-acetate. Per-acetylation of compound 3 according to

H_5III′,H_3III), 4.74 (1H, d, J ) 3.3 Hz, H_1IV), 4.62-4.48 (2H, m,

H_5I, H_4III), 4.46-4.35 (2H, m, H_2I,H_2IV), 4.25 (1H, dd, J ) 6, 6

Hz, H_5IV), 4.12-3.91 (4H, br m, H_1II, H_3II, H_4II, H_5II), 3.70-3.55

(4H, H_6IV, H_6IV′, H_6I, H_6I′), 3.48 (1H, dd, J ) 5.2, 4 Hz, H_3IV), 3.32

(1H, d, J ) 4 Hz, H_4IV), 2.48 (3H, s, -OCOCH₃), 2.42 (3H, s,

-OCOCH₃), 2.30 (1H, m, H_2IIeq), 2.09 (3H, s, -OCOCH₃), 2.05

(3H, s, -OCOCH₃), 1.96 (1H, m, H_2IIax), 1.67 (9H, s, -COC(CH₃)₃),

1.65 (9H, s, -COC(CH₃)₃), 1.58 (27H, m, 3 × -COC(CH₃)₃), 1.56

(9H, s, -COC(CH₃)_3.¹³C {¹H} NMR (100 MHz, pyridine-d₅, 363

K): δ 131.9 (ArC_AA′), 124.3 (ArC_XX′), 109.3 (C_1III), 99.6 (C_1V),

98.2 (C_1I), 84.3 (C_5II/C_4II), 80.0 (C_4III), 79.9 (C_5II/C_4II), 77.0 (C_6II),

75.8 (C_3III), 75.3 (C_2III), 73.2 (C_3I), 72.7 (C_5IV), 71.1 (C_4I), 69.9

(C_5I), 65.6 (C_5III), 54.5 (C_2I), 52.5 (C_4IV), 52.3 (C_3IV), 50.8 (C_1II,

C_6II), 46.1 (C_2IV), 43.0-42.0 (C_6I, C_6IV), 35.4 (C_2II), 30-27 (6 ×

Boc (CH₃)₃), 22.0-19.0 (4 × OAc CH₃). LCMS (ESI): calculated

for (M + Na⁺) C₆₈H₁₀₃N₇O₃₁1536.66, found 1536.7.

1

the general procedure afforded 3-penta-O-acetate. H NMR (400

MHz, pyridine-d₅, 363 K): δ 6.96 (1H, d, J ) 9 Hz, NH_II), 6.77

(2H, br m, NH_II, NH_IV), 6.57 (1H, br dd, NH_I), 6.46 (1H, d, J )

10 Hz, NH_I), 6.16 (1H, d, J ) 9.3 Hz, NH_IV), 6.02 (1H, br s, H_1I),

5.63 (1H, dd, J ) 9.5, 10.6 Hz, H_3I), 5.57 (1H, d, J ) 2.0 Hz,

H

_1III), 5.40 (1H, dd, J ) 9.5, 10 Hz, H_4I), 5.35 (1H, dd, J ) 2.4,

5.3 Hz, H_2III), 5.21 (1H, dd, J ) 10.2, 9.4 Hz, H_6II), 4.71 (1H, m,

_{5 III}), 4.68 (1H, d, J ) 3.4 Hz, H_1IV), 4.66-4.55 (3H, br m, H_3III

H

,

H_5I, H_5III), 4.46 (1H, ddd, J ) 10.4,10.4, 4.3 Hz, H_2I), 4.38 (2H,

m, H_2IV, H_4III), 4.22 (1H, dd, J ) 6, 6 Hz, H_5IV) 4.14-3.9 (4H, br

m, H_1II, H_3II, H_4II, H_5II), 3.70 (4H, H_6IV, H_6IV’, H_6I, H_6I’) 3.48 (1H,

dd, J ) 5.4, 3.8 Hz, H_3IV), 3.30 (1H, d, J ) 3.8 Hz, H_4IV), 2.45

(3H, s, -OCOCH₃), 2.40 (3H, s, -OCOCH₃), 2.40 (1H, m, H_2IIeq),

2.28 (3H, s, -OCOCH₃), 2.15 (3H, s, -OCOCH₃), 2.09 (3H, s,

-OCOCH₃), 2.0 (1H, m, H_2IIax), 1.68 (9H, s, -COC(CH₃)₃), 1.64

(9H, s, -COC(CH₃)₃), 1.60 (9H, s, -COC(CH₃)₃), 1.58 (18H, s,

2 × -COC(CH₃)₃), 1.56 (9H, s, -COC(CH₃)₃). ¹³C {¹H} NMR (100

MHz, pyridine-d₅, 363 K): δ 171.3-170.0⁴³(5 × OAc CdO),

157.2-156.3⁴³(6 × Boc CdO), 108.8 (C_1III), 99.8 (C_1IV), 98.6 (C_1I),

83.3 (C_5II/C_4II), 80.2 (C_5II/C_4II), 80.4 (C_4III), 79.3-79.0⁴³(6 × Boc

C_q), 77.5 (C_6II), 75.8 (C_3III, C_2III), 73.4 (C_3I), 73.0 (C_5IV), 71.4 (C_4I),

70.2 (C_5I), 64.1 (C_5III),), 54.8 (C_2I), 52.7 (C_4IV), 52.5 (C_3IV), 51.1

(C_1II, C_3II), 46.4 (C_2IV), 43.7 (C_6IV), 42.3 (C_6I), 35.8 (C_2II), 29.2-

3′′′-Azido-1,3,2′,6′,2′′′,6′′′-hexa-N-(tert-butoxycarbonyl)-3′′′-

deoxy-neomycin B (6).^18aNaN₃(130 mg, 2 mmol) was added to

a solution of 3 (400 mg, 0.33 mmol) in DMF (10 mL) and the

mixture was stirred at room temperature (2 d). The reaction mixture

was then diluted with ethyl acetate (200 mL) and washed with water

(2 × 100 mL). The ethyl acetate layer was dried (Na₂SO₄),

concentrated in vacuo, and purified by flash chromatography (DCM/

MeOH 97:3) yielding 6 as a white amorphous solid (352 mg, 85%).

1974 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

IR (KBr disk) 2109.9 cm^-1(N₃). HRMS (ESI): calculated for (M

+ Na⁺) C₅₃H₉₃N₉O₂₄1262.62311, found 1262.62303.

m, H_2IV, H_5III, H_3III), 4.63 (1H, dd, J ) 11.8, 5 Hz, H_5III′), 4.60

(1H, ddd, J ) 4, 4, 10 Hz, H_5I), 4.54 (1H, ddd, J ) 3.8, 5.6, 5.6

Hz, H_4III), 4.46 (1H, ddd, J ) 3.5, 10, 10 Hz, H_2I), 4.13-3.91 (5H,

m, H_1II,H_3IIH_4II,H_5II,H_6IV), 3.71 (2H, m, H_6I,H_6I′), 3.67 (1H, m,

6-Hexa-O-acetate. Per-acetylation of 6, using the general

procedure above, afforded 6-hexa-O-acetate as a white amorphous

1

solid in quantitative yield. H NMR (400 MHz, pyridine-d₅, 363

H_6IV′), 2.39 (3H, s, -OCOCH₃), 2.37 (1H, m, H_2IIeq), 2.28 (3H, s,

K): δ 6.95 (1H, d, J ) 8 Hz, NH_II), 6.78 (2H, br dd, NH_II, NH_IV),

6.58 (1H, br dd, NH_I), 6.43 (2H, m, NH_I, NH_IV), 5.97 (1H, br s,

H_1I), 5.63 (1H, dd, J ) 9.5, 10.7 Hz, H_3I), 5.59 (1H, d, J ) 2.8 Hz,

-OCOCH₃), 2.27 (3H, s, -OCOCH₃), 2.14 (6H, m, 2 × -OCOCH₃),

2.10 (3H, s, -OCOCH₃), 2.0 (1H, m, H_2IIax), 1.67 (9H, s, -COC-

(CH₃)₃), 1.62 (9H, s, -COC(CH₃)₃), 1.58 (9H, s, -COC(CH₃)₃), 1.55

(18H, s, 2 × -COC(CH₃)₃), 1.51 (9H, s, -COC(CH₃)₃). ¹³C {¹H}

NMR (100 MHz, pyridine-d₅, 363 K): δ 171.4-170.0⁴³(6 × OAc

CdO), 165.0⁴³(Ar_ipso), 157.4-156.4⁴³(6 × Boc CdO), 151.34⁴³

(Ar_ipso), 143.15⁴³(triazole_ipso), 131.3 (ArC_AA′), 126.27 (CH, triazole),

124.0 (ArC_XX′), 108.1 (C_1III), 99.0 (C_1IV), 98.7 (C_1I), 83.5 (C_5II),

80.4 (C_4II,C_4III), 79.9-78.8⁴³(6 × Boc C_q), 77.0 (C_6II), 76.5 (C_3III),

75.4 (C_2III), 73.8 (C_5IV), 73.3 (C_3I), 71.5 (C_4I), 70.2 (C_5I), 69.9 (C_4IV),

64.4 (C_5III), 61.7 (C_3IV), 59.6 (OCH₂, linker), 54.7 (C_2I), 52.5 (C_2IV),

50.9 (C_1II, C_3II), 42.3 (C_6I), 41.9 (C_6IV), 35.5 (C_2II), 29.4-28.4⁴³

(6 × Boc (CH₃)₃), 22.0-21.0⁴³(6 × OAc CH₃). HRMS (ESI):

calculated for (M + Na⁺) C₇₅H₁₁₂N₁₀O₃₄1719.72401, found

1719.72346.

H_1III), 5.40 (2H, m, H_4I, H_2III), 5.22 (1H, dd, J ) 10, 9 Hz, H_6II),

5.14 (1H, d, J ) 2.3 Hz, H_1IV), 5.13 (1H, ddd, J ) 3, 2, 1 Hz,

H

_4IV), 4.73 (2H, m, H_5III, H_3III), 4.63 (1H, dd, J ) 11.8, 5.2 Hz,

_5III′), 4.59 (1H, ddd, J ) 4, 4, 10 Hz, H_5I), 4.49-4.42 (3H, br m,

H_2I, H_3IV, H_4III), 4.37 (1H, ddd, J ) 2.6, 5, 7 Hz, H_5IV), 4.24 (1H,

br m, H_2IV), 4.10 (1H, dd, J ) 8.5,10 Hz, H_4II), 4.07-3.97 (2H, br

m, H_1II,H_3II), 3.97 (1H, dd, J ) 9, 9 Hz, H_5II), 3.83 (1H, ddd, J )

7, 7, 14 Hz, H_6IV), 3.71 (2H, m, H_6I, H_6I′), 3.60 (1H, ddd, J ) 5,

6.5, 14 Hz, H_6IV′), 2.39 (3H, s, -OCOCH₃), 2.39 (1H, m, H_2IIeq),

2.34 (3H, s, -OCOCH₃), 2.28 (3H, s, -OCOCH₃), 2.14 (3H, s,

-OCOCH₃), 2.13 (3H, s, -OCOCH₃), 2.09 (3H, s, -OCOCH₃), 2.01

(1H, m, H_2IIax), 1.67 (9H, s, -COC(CH₃)₃), 1.61 (9H, s, -COC-

(CH₃)₃), 1.58 (18H, s, 2 × -COC(CH₃)₃), 1.57 (9H, s, -COC(CH₃)₃),

1.56 (9H, s, -COC(CH₃)₃). ¹³C {¹H} NMR (100 MHz, pyridine-d₅,

363 K): δ 171.3-170.2⁴³(6 × OAc CdO), 157.4-156.4⁴³(6 ×

Boc CdO), 107.9 (C_1III), 98.9 (C_1IV), 98.7 (C_1I), 83.6 (C_5II), 80.4

(C_4II), 80.3 (C_4III), 79.8-78.8⁴³(6 × Boc C_q), 77.2 (C_6II), 76.9 (C_3III),

75.5 (C_2III), 73.3 (C_5IV, C_3I), 71.3 (C_4I), 70.0 (C_5I), 68.0 (C_4IV), 64.1

(C_5III), 60.8 (C_3IV), 54.7 (C_2I), 51.3 (C_2IV), 50.9 (C_1II, C_3II), 42.2

(C_6I), 41.6 (C_6IV), 35.4 (C_2II), 29.1-28.9⁴³(6 × Boc (CH₃)₃), 21.9-

21.1⁴³(6 × OAc CH₃). HRMS (ESI): calculated for (M + Na⁺)

C₆₅H₁₀₅N₉O₃₀1514.68650, found 1514.68447.

Prop-2-ynyl Dodecanate (9).⁴⁵Propargyl alcohol (60 µL, 1

mmol) was added to a stirred solution of lauric acid (100 mg, 0.5

mmol) and DCC (120 mg, 0.58 mmol) in dry DCM (3 mL), and

the resultant mixture was stirred at room temperature (4 h). The

reaction mixture was then diluted with acetone and filtered. The

filtrate was concentrated, diluted with ethyl acetate (50 mL), and

washed with saturated aqueous Na₂CO₃solution followed by water

(3 × 15 mL).The organic phase was dried (Na₂SO₄) and evaporated,

leaving prop-2-ynyl dodecanate 9 as a light yellow oil (110 mg,

92%). The crude product was found to be of sufficient purity for

use in the next step without need for further purification. ¹H NMR

(400 MHz, CDCl_3,298 K): δ 4.65 (2H, d, J ) 2.5 Hz) 2.44 (1H,

t, J ) 2.5 Hz), 2.36 (2H, t, J ) 7.5 Hz), 1.62 (2H, m), 1.34-1.18

(16H, m), 0.86 (3H, m). ¹³C {¹H} NMR (100 MHz, CDCl₃, 298

K): δ 173.2 (CdO), 78.0 (q C, -CtCH), 74.9(CH), 52.0 (-OCH₂),

34.2, 32.1, 29.8-29.3, 25.0, 22.9, 14.3.

Prop-2-ynyl 4-Nitrobenzoate (7). Propargyl alcohol (50 µL,

0.86 mmol) was added to a stirred solution of 4-nitrobenzoyl

chloride (100 mg, 0.5 mmol) in pyridine (1 mL) and the solution

was stirred at room temperature (4 h). Aqueous HCl acid solution

(1 N, 12 mL) was added to the reaction mixture and the resultant

mixture was extracted with ethyl acetate (50 mL). The organic layer

was washed with Na₂CO_3(sat)solution (15 mL) and water (3 × 15

mL) and then dried (Na₂SO₄). Concentration in vacuo afforded prop-

2-ynyl 4-nitrobenzoate 7 (105 mg, 94%) as a brown solid which,

in general, was used in succeeding steps without further purification.

Recrystallization of the crude product from hexane afforded 7 as

fine white needles, mp 90-92 °C {lit.⁴⁴mp 89-90 °C}. ¹H NMR

(400 MHz, CDCl_3,298 K): δ 8.36 (4H, m), 4.95 (2H, d, J ) 2.5

Hz) 2.49 (1H, t, J ) 2.5 Hz). ¹³C {¹H} NMR (100 MHz, CDCl₃,

298 K): δ 164.5 (CdO), 150.6 (ipso), 135.0 (ipso), 131.2 (Ar),

123.8 (Ar), 77.2(Cq), 75.9 (CH), 53.5 (-OCH₂).

1,3,2′,6′,2′′′,6′′′-Hexa-N-(tert-butoxycarbonyl)-3′′′-deoxy-3′′′-

(4-dodecanoyloxymethyl-[1,2,3]-triazol-1-yl)-neomycin B (13).

A mixture of tert-butanol and water (1:1, 2 mL) was added to a

mixture of the 3′′′-azidoneomycin 6 (50 mg 0.04 mmol), prop-2-

ynyl dodecanate 9 (12 mg, 0.05 mmol), CuSO₄·5H₂O (1 mg), and

copper powder (30 mg). The reaction mixture was stirred vigorously

at room temperature (18 h) and then diluted with ethyl acetate (20

mL). Copper powder was removed by filtration and the filtrate was

dried (Na₂SO₄) and concentrated in vacuo. Purification by flash

chromatography on silica (DCM/MeOH 95:5) afforded 13 (43 mg,

72%) as an amorphous white solid. HRMS (ESI): calculated for

(M + Na⁺) C₆₈H₁₁₉N₉O₂₆1500.81639, found 1500.81614.

13-Hexa-O-acetate. Per-acetylation of compound 13 using the

general method above afforded 13-hexa-O-acetate as an amorphous

white solid. ¹H NMR (400 MHz, pyridine-d₅, 363 K): δ 8.25 (1H,

s, ArH, triazole), 6.94 (1H, d, J ) 8.5 Hz, NH_IV), 6.78 (3H, m, 2

× NH_II, NH_IV), 6.62 (1H, br dd, NH_I), 6.46 (1H, d, J ) 9.6 Hz,

NH_I), 5.98 (1H, br s, H_1I), 5.71 (1H, dd, J ) 3, 3 Hz, H_4IV), 5.64

(1H, dd, J ) 9.3, 10.5 Hz, H_3I), 5.61 (1H, d, J ) 2.4 Hz, H_1III),

5.58 (1H, d, J ) 2.7 Hz, H_1IV), 5.45 (1H, dd, J ) 2.6, 5 Hz, H_2III),

5.43 (2H, s, OCH₂), 5.40 (1H, dd, J ) 9.6, 9.6 Hz, H_4I), 5.38 (1H,

dd, J ) 3.5, 5.7 Hz, H_3IV), 5.21 (1H, dd, J ) 9.5, 10.4 Hz, H_6II),

4.88 (1H, ddd, J ) 2.5, 6.5, 6.5 Hz, H_5IV), 4.82-4.77 (3H, m, H_2IV,

H_5III,H_3III), 4.64 (1H, dd, J ) 11.8, 5.3 Hz, H_5III′), 4.59 (1H, ddd,

J ) 4, 4, 10 Hz, H_5I), 4.54 (1H, ddd, J ) 3.8, 5.6, 5.6 Hz, H_4III),

4.46 (1H, ddd, J ) 3.7, 10.3, 10.3 Hz, H_2I), 4.12-3.88 (5H, m,

H_1II, H_3II, H_4II, H_5II, H_6IV), 3.71 (2H, m, H_6I, H_6I′), 3.67 (1H, m,

6,3′,4′,2′′,5′′,4′′′-Hexa-O-acetyl-1,3,2′,6′,2′′′,6′′′-hexa-N-(tert-bu-

toxycarbonyl)-3′′′-deoxy-3′′′-[4-(4-nitrobenzoyloxymethyl)-[1,2,3]-

triazol-1-yl)-neomycin B (8-hexa-O-acetate). A mixture of tert-

butanol and water (1:1, 2 mL) was added to the mixture of 6-hexa-

O-acetate (50 mg 0.04 mmol), prop-2-ynyl 4-nitrobenzoate, 7 (10

mg, 0.05 mmol), CuSO₄·5H₂O (1 mg), and copper powder (30 mg).

The resultant reaction mixture was stirred vigorously at room

temperature (18 h) and then diluted with ethyl acetate (20 mL).

Residual copper powder was removed by filtration and the filtrate

was dried (Na₂SO₄) and concentrated in vacuo. Purification by flash

chromatography on silica (DCM/MeOH 98:2) afforded 8-hexa-O-

acetate (46 mg, 79%) as an amorphous white solid. ¹H NMR (400

MHz, pyridine-d₅, 363 K): δ 8.40 (1H, s, ArH, triazole), 8.28 (2H,

m, ArH_XX′), 8.21 (2H, m, ArH_AA′), 6.94 (1H, d, J ) 8.5 Hz, NH_IV),

6.79 (3H, m, 2 × NH_II, NH_IV), 6.61 (1H, br dd, NH_I), 6.40 (1H, d,

J ) 10 Hz, NH_I), 5.98 (1H, br s, H_1I), 5.75 (1H, dd, J ) 3, 3 Hz,

H_4IV), 5.67 (2H, s,OCH₂), 5.63 (1H, dd, J ) 9.5, 10.5 Hz, H_3I),

5.60 (1H, d, J ) 2.5 Hz, H_1III), 5.57 (1H, d, J ) 3 Hz, H_1IV), 5.44

(1H, dd, J ) 2.5, 5 Hz, H_2III), 5.41 (1H, dd, J ) 3.5, 6 Hz, H_3IV),

5.40 (1H, dd, J ) 9.5, 9.5 Hz, H_4I), 5.21 (1H, dd, J ) 10, 10 Hz,

H_6II), 4.90 (1H, ddd, J ) 2.5, 6.7, 6.7 Hz, H_5IV), 4.83-4.75 (3H,

H

_6IV′), 2.41 (2H, t, J ) 7.4 Hz, -COCH₂, H_D^/), 2.40 (3H, s,

-OCOCH₃), 2.38 (1H, m, H_2IIeq), 2.32 (3H, s, -OCOCH₃), 2.28

(3H, s, -OCOCH₃), 2.15 (6H, m, 2 × -OCOCH₃), 2.10 (3H, s,

-OCOCH₃), 2.0 (1H, m, H_2IIax), 1.74 (2H, m, H_D^/), 1.67 (9H, s,

(44) Reppe, W. Justus Liebigs Ann. Chem. 1955, 596, 38-79.

(45) Masaki, Y.; Tanaka, N.; Miura, T. Chem. Lett. 1997, 55-56.

J. Org. Chem, Vol. 72, No. 6, 2007 1975

Quader et al.

-COC(CH₃)₃), 1.62 (9H, s, -COC(CH₃)₃), 1.58 (9H, s, -COC(CH₃)₃),

1.56 (9H, s, -COC(CH₃)₃), 1.55 (9H, s, -COC(CH₃)₃), 1.51 (9H, s,

-COC(CH₃)₃), 1.47-1.32 (16H, m, H_D^/), 0.97 (3H, distorted t, H_D^/).

¹³C {¹H} NMR (100 MHz, pyridine-d₅, 363 K): δ 125.25 (CH,

75%). HRMS (ESI): calculated for (M + Na⁺) C₇₇H₁₂₅N₉O₂₇

1630.85826, found 1630.86152.

14-Hepta-O-acetate. Per-acetylation of compound 14 according

to the general procedure above afforded 14-hexa-O-acetate as an

1

triazole), 107.8 (C_1III), 98.8 (C_1IV), 98.4 (C_1I), 83.3 (C_5II), 80.3 (C_4III

)

amorphous white solid in quantitative yield. H NMR (400 MHz,

,80.0 (C_4II), 76.7 (C_6II), 76.4 (C_3III), 75.22 (C_2III), 75.5 (C_5IV), 73.1

(C_3I), 71.1 (C_4I), 69.9 (C_5I), 69.8 (C_4IV), 64.1 (C_5III), 61.3 (C_3IV),

58.0 (OCH₂, linker), 54.5 (C_2I), 52.2 (C_2IV), 50.8 (C_1II, C_3II), 42.0

(C_6I), 41.6 (C_6IV), 35.7 (C_2II), 34.4 (C_D^/, -CH₂-CO-), 25.5 (C_D^/),

30.0-27.0 (6 × Boc (CH₃)₃), 22.7-20.0 (6 × OAc CH₃), 14.0

pyridine-d₅, 363 K): δ 8.29 (1H, s, ArH, triazole), 6.95 (1H, d,

J ) 8.4 Hz, NH_IV), 6.84-6.74 (3H, m, 2 × NH_II, NH_IV), 6.63 (1H,

br dd, NH_I), 6.46 (1H, d, J ) 10.5 Hz, NH_I), 5.98 (1H, br s, H_1I),

5.77(1H, t, J ) 2.5 Hz ,H_21S^/), 5.72 (1H, dd, J ) 3, 3 Hz, H_4IV),

5.64 (1H, dd, J ) 9.4, 10.7 Hz, H_3I), 5.62 (1H, d, J ) 2.4 Hz,

(C_D^/, -CH₃). HRMS (ESI): calculated for (M + Na⁺) C₈₀H₁₃₁N₉O₃₂

H_1III), 5.60 (1H, d, J ) 2.7 Hz, H_1IV), 5.50 (2H, s, OCH₂), 5.45-

/

(1H, m, H_6S^/), 5.43 (1H, m, H_2III), 5.41 (1H, dd, J ) 9.7, 9.7 Hz,

H_4I), 5.39 (1H, dd, J ) 3.5, 5.5 Hz, H_3IV), 5.22 (1H, dd, J ) 9.4,

10.2 Hz, H_6II), 4.89 (1H, ddd, J ) 2.6, 6.7, 6.7 Hz, H_5IV), 4.85-

4.76 (4H, m, H_2IV, H_5III,H_3III,H_3S^/), 4.65 (1H, dd, J ) 11.7, 5.0

Hz, H_5III′), 4.62 (1H, ddd, J ) 3.9, 3.9, 8.0 Hz, H_5I), 4.55 (1H,

ddd, J ) 3.8, 5.3, 5.3 Hz, H_4III), 4.47 (1H, ddd, J ) 4.0, 10.5, 10.5

Hz, H_2I), 4.14-3.88 (5H, m, H_1II,H_3IIH_4II,H_5II,H_6IV), 3.71 (2H,

m, H_6I,H_6I′), 3.68 (1H, m, H_6IV′),3.03 (2H, m, H_16a,bS^/), 2.53 (2H,

m, H_4a,bS), 2.40 (3H, s, -OCOCH₃), 2.38 (1H, m, H_2IIeq), 2.31

(3H, s, -OCOCH₃), 2.28 (3H, s, -OCOCH₃), 2.14 (6H, m, 2 ×

-OCOCH₃), 2.10 (6H, s, 2 × -OCOCH₃), 2.0 (1H, m, H_2IIax), 1.67

1752.87978, found 1752.87986. ) dodecanoyl

D

3â-Hydroxypregna-5,17(20)-diene-21-oic Acid (12).⁴⁶Pow-

dered KOH (70 mg) was added to a stirred solution of ethyl 3â-

hydroxypregna-5,17(20)-diene-21-oate 11 (200 mg, 0.55 mmol) in

isopropanol (3 mL) and the reaction mixture was refluxed for 2 h.

The mixture was then cooled to room temperature and evaporated

to dryness. The residue was partitioned between ethyl acetate and

1 M aqueous HCl acid solution. The ethyl acetate layer was washed

with water, dried (Na₂SO₄), and concentrated in vacuo. Flash

column chromatography (DCM/MeOH 98:2) afforded product 3â-

hydroxypregna-5,17(20)-diene-21-oic acid 12 (130 mg, 72%) as a

white crystalline solid, mp 253-255 °C {lit.⁴⁶mp 252-254 °C}.

¹H NMR (400 MHz, pyridine-d₅, 298 K): δ 6.00 (1H, dd, J ) 2.5,

2.5 Hz, H20), 5.43 (1H, m, H6), 3.86 (1H, dddd, J ) 4.1, 6.1,

10.5, 10.5 Hz, H3), 3.18-3.12 (2H, m, H16a,b), 2.64-2.58 (2H,

m, H4a,b), 2.11 (1H, m, H2a), 2.01 (1H, m, H7a), 1.88-1.78 (3H,

br m, H12a, H1a, H2b), 1.73 (1H, m, H15a), 1.65-1.54 (2H, br

m, H11a, H7b), 1.54-1.40 (2H, br m, H8, H11b), 1.35-1.23 (2H,

br m, H15b, H12b), 1.12 (1H, ,H1b), 1.06-0.95 (2H, m, H14, H9),

1.06 (3H, s, CH₃, H19), 0.81 (3H, s, CH₃, H18). ¹³C{¹H} NMR

(100 MHz, pyridine-d₅, 298 K): δ 175.5 (C17), 170.3 (CdO, C21),

142.5 (C5), 121.5 (C6), 111.0 (C20), 71.7 (C3), 54.6 (C14), 51.1

(C9), 46.6 (C13), 44.0 (C4), 38.3 (C1), 37.5 (C10), 36.1 (C12),

33.1 (C2), 32.5 (C7), 32.4 (C8), 31.3 (C16), 25.2 (C15), 21.8(C11),

20.1 (C19), 18.9 (C18).

(9H, s, -COC(CH₃)₃), 1.62 (9H, s, -COC(CH₃)₃), 1.58 (9H, s, -COC-

/

(CH₃)₃), 1.57-1.53 (27H, m, 3 × -COC(CH₃)₃), 1.10 (3H, s, H_19S

,

CH₃), 0.88 (3H, s,H_18S^/, CH₃). ¹³C {¹H} NMR (100 MHz, pyridine-

d₅, 363 K): δ 177.6⁴³(C_17S^/), 171.4-170.1⁴³(7 × OAc CdO),

167.2 (CdO, C_21S^/) ,157.5-156.3⁴³(6 × Boc CdO), 144.1⁴³(ipso,

triazole), 140.5⁴³(C_5S^/), 125.3 (CH, triazole), 122.7 (C_6S^/) 109.3

(C_20S^/) 108.0 (C_1III), 99.0 (C_1IV), 98.6 (C_1I), 83.4 (C_5II), 80.5 (C_4III

)

,80.3 (C_4II), 80-78.7⁴³(6 × Boc C_q), 77.0 (C_6II), 76.6 (C_3III), 75.5

(C_2III), 74.4 (C_3S^/), 73.3 (C_3I), 72.6 (C_5IV), 71.2 (C_4I), 71.1 (C_5I),

70.0 (C_4IV), 64.4 (C_5III), 61.5 (C_3IV), 57.7 (OCH₂, linker), 54.8 (C_2I),

52.6 (C_2IV) 52.0-48.0 (C_1II, C_3II), 41.9 (C_6I), 41.73 (C_6IV), 38.7

(C_4S^/), 35.67 (C_2II), 31.2 (C_16S^/), 29.3-28.7⁴³(6 × Boc (CH₃)₃),

21.8-21.0⁴³(7 × OAc CH₃), 19.77(C_19S^/), 18.58(C_18S^/). HRMS

(ESI): calculated for (M + Na⁺) C₉₁H₁₃₉N₉O₃₄1924.93221, found

/

1924.93161. ) steroid

Prop-2-ynyl 3â-Hydroxypregna-5,17 (20)-diene-21-oate (10).

DIAD (65 µL, 0.34 mmol) was added slowly to a stirred solution

of 3â-hydroxypregna-5,17(20)-diene-21-oic acid 12 (75 mg, 0.23

mmol), TPP (90 mg, 0.34 mmol), and anhydrous propargyl alcohol

(30 µL, 0.44 mmol) in dry THF (1 mL) under N₂atmosphere and

the mixture was stirred at room temperature (12 h). The solvent

was removed in vacuo and the residue was purified by column

chromatography (DCM/MeCN 95:5) affording ester 10 (65 mg,

75%) as white solid, mp 158-159 °C . ¹H NMR (400 MHz, CDCl₃,

298 K): δ 5.61 (1H, dd, J ) 2.5, 2.5 Hz, H20), 5.37 (1H, m, H6),

4.71 (2H, d, J ) 2.5, OCH₂, H1′), 3.54 (1H, dddd, J ) 4, 5, 11, 11

Hz, H3), 2.90-2.83 (2H, m, H16a,b), 2.47 (1H, t, J ) 2.5 Hz,

H3′) 2.36-2.20 (2H, m, H4a,b), 2.03 (1H, m, H7a), 1.92-1.77

(4H, br m, H12a, H15a, H1a, H2a), 1.74-1.46 (5H, br m, H11a,b,

H8, H2b, H7b), 1.44-1.22 (2H, br m, H15b, H12b), 1.16-0.96

(3H, m, H1b, H14, H9), 1.0 (3H, s, CH₃, H19), 0.85 (3H, s, CH₃,

H18); ¹³C{¹H} NMR (100 MHz, CDCl₃, 298 K): δ 178.5 (C17),

166.5 (CdO, C21), 141.0 (C5), 121.5 (C6), 107.8 (C20), 78.5 (C

2′), 74.6 (C3′), 71.9 (C3), 54.0 (C14), 51.4 (C1′), 50.4 (C9), 46.5

(C13), 42.4 (C4), 37.4 (C1), 36.8 (C10), 35.3 (C12), 31.9-31.8

(C7, C2, C8), 30.8 (C16), 24.7 (C15), 21.2 (C11), 19.6 (C19), 18.4

(C18). HRMS (EI): calculated for (M⁺) C₂₄H₃₂O₃368.23514, found

368.23541.

S

1,3,2′,6′,2′′′,6′′-Hexa-N-(tert-butoxycarbonyl)-5′′-O-(2,4,6-tri-

isopropylbenzenesulfonyl)-neomycin B (15).³⁰A solution of 2 (1

g, 0.82 mmol) and excess 2,4,6-triisopropylbenzenesulfonyl chloride

(7 g, 23 mmol) in dry pyridine (20 mL) was stirred at room

temperature (18 h). Pyridine was removed in vacuo by coevapo-

ration with toluene. The crude residue was then dissolved in ethyl

acetate (200 mL) and washed with water (2 × 100 mL). The

aqueous layers were combined and extracted with ethyl acetate

(2 × 100 mL). The combined organic layers were dried (Na₂SO₄),

concentrated in vacuo, and subjected to flash chromatography.

Unreacted 2,4,6-triisopropylbenzenesulfonyl chloride (6 g, mp

identical with that of authentic sample) was recovered using DCM

as eluent. The residue (1.2 g) was eluted from the silica column

(DCM/MeOH 5:1) and then rechromatographed (DCM/MeOH 98:2

f 95:5) affording 15 as an amorphous white solid (0.75 g, 61%).

¹H NMR (400 MHz, methanol-d₄, 298 K): δ 7.33 (s, 2H), 5.52

(br s, 1H), 5.22 (br s, 1H), 4.99 (br s, 1H), 4.39 (m, 1H), 4.30 (m,

2H), 4.19 (m, 4H), 3.91 (m, 1H), 3.82 (m, 1H), 3.77 (m, 2H), 3.62

(m, 1H), 3.54 (m, 4H), 3.47-3.36 (m, 4H), 3.24 (m, 2H), 2.96 (m,

2H), 1.96 (m, 1H), 1.51-1.40 (m, 54H), 1.31 (m, 18H). LRMS

(ESI): calculated for (M + Na⁺) C₆₈H₁₁₆N₆O₂₇S 1504, found 1504.

5′′-Azido-1,3,2′,6′,2′′′,6′′′-hexa-N-(tert-butoxycarbonyl)-5′′-

deoxy-neomycin B (16). A solution of 15 (500 mg, 0.3 mmol)

and NaN₃(150 mg, 2.3 mmol) in DMF (5 mL) was stirred at

elevated temperature (100 °C, 8 h). After cooling to room

temperature the reaction mixture was diluted with ethyl acetate (100

mL) and washed with water (2 × 20 mL). The ethyl acetate layer

was dried (Na₂SO₄) and evaporated, yielding 16 (365 mg, 98%) as

a white amorphous solid. The crude product was of sufficient purity

to use in the next step without need for further purification. IR

(KBr disk) 2106.3 cm^-1(N₃). HRMS (ESI): calculated for (M +

Na⁺) C₅₃H₉₃N₉O₂₄1262.62311, found 1262.62835.

1,3,2′,6′,2′′′,6′′′-Hexa-N-(tert-butoxycarbonyl)-3′′′-deoxy-3′′′-

[4-(3â-hydroxypregna-5,17(20)-diene-21-oyloxymethyl)- [1,2,3]-

triazol-1-yl]-neomycin B (14). The 3′′′-azidoneomycin 6 (50 mg

0.04 mmol) was treated with ester 10 (18 mg, 0.05 mmol), according

to the procedure described for the preparation of compound 13.

Purification by flash chromatography on silica (DCM/MeOH 97:

3) afforded the product 14 as an amorphous white solid (48 mg,

(46) Sondheimer, F.; Mancera, O.; Urquiza, M.; Rosenkranz, G. J. Am.

Chem. Soc. 1955, 77, 4145-4149.

1976 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

16-Hexa-O-acetate. Per-acetylation of the 5′′-azido neomycin

B 16 according to the general method afforded 16-hexa-O-acetate

as an amorphous white solid, in quantitative yield. H NMR (400

HRMS (ESI): calculated for (M + H⁺) C₇₅H₁₁₂N₁₀O₃₄1697.74207,

found 1697.74399.

1

1,3,2′,6′,2′′′,6′′′-Hexa-N-(tert-butoxycarbonyl)-5′′-deoxy-5′′(4-

dodecanoyloxymethyl-[1,2,3]triazol-1-yl)-neomycin B (18). 5′′-

Azido neomycin 16 (50 mg 0.04 mmol) was treated with ester 9

as per the method described for the synthesis of compound 13.

Chromatographic purification of the residue on silica (DCM/MeOH

96:4) afforded 18 (42 mg, 71%) as an amorphous white solid.

HRMS (ESI): calculated for (M + Na⁺) C₆₈H₁₁₉N₉O₂₆1500.81639,

found 1500.81215.

MHz, pyridine-d₅, 363 K): δ 6.96 (1H, d, J ) 8.7 Hz, NH_II), 6.80

(1H, d, J ) 8.7 Hz, NH_II), 6.70 (1H, br dd, NH_IV), 6.60 (1H, br dd,

NH_I), 6.49 (1H, d, J ) 10 Hz, NH_I), 6.12 (1H, d, J ) 10 Hz, NH_IV),

5.99 (1H, br s, H_1I), 5.64 (1H, dd, J ) 9.4, 10.7 Hz, H_3I), 5.61

(1H, d, J ) 2 Hz, H_1III), 5.51 (1H, dd, J ) 3, 3 Hz, H_3IV), 5.40

(1H, dd, J ) 9.5, 10.1 Hz, H_4I), 5.39 (1H, dd, J ) 2.5, 5 Hz, H_2III),

5.25 (1H, m, H_6II), 5.23 (1H, m, H_4IV), 5.18 (1H, d, J ) 2.12 Hz,

18-Hexa-O-acetate. Per-acetylation of 18, according to the

general procedure, afforded 18-hexa-O-acetate as an amorphous

white solid in quantitative yield. ¹H NMR (400 MHz, pyridine-d₅,

363 K): δ 8.34 (1H, s, ArH, triazole), 7.01 (1H, d, J ) 8.5 Hz

NH_II), 6.89 (1H, br dd, NH_IV), 6.80 (1H, d, J ) 7.7 Hz, NH_II),

6.67 (2H, m, 2 × NH_I), 6.06 (1H, d, J ) 9.8 Hz, NH_IV), 5.92 (1H,

d, J ) 3.5 Hz H_1I), 5.67 (1H, dd, J ) 9.3, 10.6 Hz, H_3I), 5.65 (2H,

s, OCH₂), 5.63 (1H, d, J ) 2.0 Hz, H_1III), 5.52 (1H, dd, J ) 3, 3

Hz, H_3IV), 5.49 (1H, dd, J ) 9.8, 9.8 Hz, H_4I), 5.45 (1H, dd, J )

2.2, 5.3 Hz, H_2III), 5.25 (1H, br dd, H_4IV), 5.20 (1H, dd, J ) 10, 10

Hz, H_6II), 5.20 (1H, d, J ) 2 Hz, H_1IV), 5.13 (1H, dd, J ) 3.8, 14.5

Hz, H_5III), 4.93 (1H, dd, J ) 6.5, 14.5 Hz, H_5III′), 4.79 (1H, dd,

J ) 6.8, 5.0 Hz, H_3III), 4.68 (1H, ddd, J ) 3.6, 3.6, 9.6 Hz, H_5I),

4.58 (2H, m, H_4III,H_2I), 4.51 (1H, ddd, J ) 1.5, 6.5, 6.5 Hz, H_5IV),

4.30 (1H, m, H_2IV), 4.14-3.9 (4H, m H_1II,H_3IIH_4II,H_5II), 3.86 (1H,

m, H_6IV), 3.75 (3H, m, H_6I, H_6I′, H_6IV′), 2.57 (2H, t, J ) 7.5 Hz,

-COCH₂, H_D),2.46 (3H, s, -OCOCH₃), 2.4 (1H, m, H_2IIeq), 2.35

(3H, s, -OCOCH₃), 2.19 (3H, s, -OCOCH₃), 2.17 (3H, s, -O-

COCH₃), 2.10 (3H, s, -OCOCH₃), 2.06 (3H, s, -OCOCH₃), 2.0 (1H,

m, H_2IIax), 1.83 (2H, m, H_D), 1.70 (9H, s, -COC(CH₃)₃), 1.61 (18H,

s, 2 × -COC(CH₃)₃), 1.58 (27H, s, 3 × -COC(CH₃)₃), 1.46-1.36

(16H, m, H_D), 0.98 (3H, distorted t, H_D). ¹³C {¹H} NMR (100 MHz,

pyridine-d₅, 363 K): δ 125.8 (CH, triazole), 108.0 (C_1III), 99.5

(C_1IV), 98.6 (C_1I), 82.9 (C_5II), 80.6 (C_4III), 80.3 (C_4II), 77.5 (C_3III),

76.8 (C_6II), 74.6 (C_2III), 74.2 (C_5IV), 73.1 (C_3I), 70.8 (C_4I), 70.3 (C_3IV),

70.3 (C_5I), 67.2 (C_4IV), 58.4 (OCH₂, linker), 54.5 (C_2I), 52.3 (C_5III),

50.9 (C_1II, C_3II), 50.6 (C_2IV), 42.1 (C_6I), 41.7 (C_6IV), 35.7 (C_2II),

34.8 (C_D,-CH₂-CO-), 25.5 (C_D) 31.0-28.0 (6 × Boc (CH₃)₃), 23.0-

19.0 (6 × OAc CH₃) ,14.0 (C_D, -CH₃). HRMS (ESI): calculated

for (M + Na⁺) C₈₀H₁₃₁N₉O₃₂1752.87978, found 1752.88177.

1,3,2′,6′,2′′′,6′′′-Hexa-N-(tert-butoxycarbonyl)-5′′-deoxy-5′′-[4-

(3â-hydroxypregna-5,17(20)-diene-21-oyl-oxymethyl)-[1,2,3]tri-

azol-1-yl]-neomycin B (19). The azido neomycin 16 (50 mg 0.04

mmol) was treated with ester 10 (18 mg, 0.05 mmol) in accordance

with the procedure described for the preparation of compound 13.

Purification by flash chromatography on silica (DCM/MeOH, 97:

3) afforded 19 (45 mg, 69%) as an amorphous white solid. HRMS

(ESI): calculated for (M + Na⁺) C₇₇H₁₂₅N₉O₂₇1630.85826, found

1630.86482.

19-Hepta-O-acetate. Per-acetylation of 19, according to the

general procedure, provided 19-hepta-O-acetate. ¹H NMR (400

MHz, pyridine-d₅, 363 K): δ 8.38 (1H, s, ArH, triazole), 7.01 (1H,

d, J ) 9.5 Hz NH_II), 6.88 (1H, br dd, NH_IV), 6.78 (1H, d, J ) 8.5

Hz, NH_II), 6.71 (1H, br dd, NH_I), 6.63 (1H, d, J ) 9 Hz, NH_I),

6.05 (1H, d, J ) 9.4 Hz, NH_IV), 5.92 (1H, d, J ) 3.5 Hz H_1I), 5.86

(1H, t, J ) 2.4, H_21S), 5.73 (2H, s, OCH₂), 5.68 (1H, dd, J ) 9.4,

10.4 Hz, H_3I), 5.64 (1H, d, J ) 2.0 Hz, H_1III), 5.52 (1H, dd, J ) 3,

3 Hz, H_3IV), 5.49 (1H, dd, J ) 10, 10 Hz, H_4I), 5.86 (1H, m, H_6S),

5.45 (1H, m, H_2III), 5.25 (1H, br dd, H_4IV), 5.20 (1H, dd, J ) 9.5,

9.5 Hz, H_6II), 5.20 (1H, d, J ) 2.3 Hz, H_1IV), 5.11 (1H, dd, J ) 4,

14.2 Hz, H_5III), 4.98 (1H, dd, J ) 5.8, 14.2 Hz, H_5III′), 4.83 (1H,

m, H_3S), 4.79 (1H, dd, J ) 6.5, 5.3 Hz, H_3III), 4.67 (1H, ddd, J )

4, 4, 10 Hz, H_5I), 4.61-4.52 (2H, m, H_4III,H_2I), 4.49 (1H, ddd,

J ) 1.6, 6.5, 6.5 Hz, H_5IV), 4.30 (1H, m, H_2IV), 4.17-3.97 (4H, m

H_1II, H_3II, H_4II,H_5II), 3.87 (1H, m, H_6IV), 3.77 (1H, m, H_6I), 3.72

(2H, m, H_6I’H_6IV′), 3.10 (2H, m, H_16abS), 2.53 (2H, m, H_4abS), 2.44

(3H, s, -OCOCH₃), 2.38 (1H, m, H_2IIeq), 2.35 (3H, s, -OCOCH₃),

2.19 (3H, s, -OCOCH₃), 2.16 (3H, s, -OCOCH₃), 2.09 (6H, s, 2 ×

-OCOCH₃), 2.06 (3H, s, -OCOCH₃), 2.0 (1H, m, H_2IIax), 1.69

H_1IV), 4.76 (1H, dd, J ) 6.6, 5.3 Hz, H_3III), 4.60 (1H, ddd, J ) 4,

4,10 Hz, H_5I), 4.51 (1H, ddd, J ) 3.8, 10.5, 10.5 Hz, H_2I), 4.46

(1H, ddd, J ) 1.8, 6.7, 6.7 Hz, H_5IV), 4.32 (1H, ddd, J ) 3.8, 4.6,

6.6 Hz, H_4III), 4.30 (1H, br m, H_2IV), 4.13 (1H, dd, J ) 8.6, 10 Hz,

H_4II), 4.10-4.01 (2H, m, H_3II, H_1II), 3.98 (1H, m, H_5II), 3.97 (1H,

dd, J ) 8.6, 8.6 Hz, H_5III), 3.82 (1H, ddd, J ) 6.6, 6.6, 13.7 Hz,

H_6IV), 3.76 (1H, dd, J ) 8.6, 8.6 Hz, H_5′III), 3.70 (2H, m, H_6I,H_6I),

3.62 (1H, ddd, J ) 4.8, 6.8, 13.7 Hz, H_6′IV), 2.40 (3H, s, -OCOCH₃),

2.40 (1H, m, H_2IIeq), 2.33 (3H, s, -OCOCH₃), 2.14 (3H, s,

-OCOCH₃), 2.13 (3H, s, -OCOCH₃), 2.08 (3H, s, -OCOCH₃), 2.04

(3H, s, -OCOCH₃), 2.0 (1H, m, H_2IIax), 1.66 (9H, s, -COC(CH₃)₃),

1.62 (9H, s, -COC(CH₃)₃), 1.57 (18H, s, 2 × -COC(CH₃)₃), 1.57

(9H, s, -COC(CH₃)₃), 1.55 (9H, s, -COC(CH₃)₃). ¹³C {¹H} NMR

(100 MHz, pyridine-d₅, 298 K): δ 171.5-169.2 (6 × OAc CdO),

157.6-156.6 (6 × Boc CdO), 108.4 (C_1III), 99.0 (C_1IV), 98.4 (C_1I),

83.5 (C_5II/C_4II), 80.2 (C_4II/C_5II), 79.5 (C_4III), 79.0-78.7 (6 × Boc

C_q), 77.3 (C_6II), 76.1 (C_3III), 74.9 (C_2III), 73.8 (C_5IV), 73.3 (C_3I),

70.5 (C_4I), 69.9 (C_3IV), 69.8 (C_5I), 66.8 (C_4IV), 54.1 (C_2I), 51.9 (C_5III),

50.2 (C_2IV, C_1II, C_3II), 41.3 (C_6I), 40.8 (C_6IV), 35.9 (C_2II), 29.5-

28.5 (6 × Boc (CH₃)₃), 22.2-21.0⁴³(6 × OAc CH₃). LRMS

(ESI): calculated for (M + Na⁺) C₆₅H₁₀₅N₉O₃₀1514.7, found

1514.7.

1,3,2′,6′,2′′′,6′′′-Hexa-N-(tert-butoxycarbonyl)-5′′-deoxy-5′′-[4-

(4-nitrobenzoyloxymethyl)-[1,2,3]triazol-1-yl]-neomycin B (17).

The synthetic procedure for compound 13 was repeated using the

5′′-azido neomycin 16 (50 mg 0.04 mmol) and ester 7 (10 mg,

0.05 mmol). Purification of the crude product by flash chromatog-

raphy on silica (DCM/MeOH 97:3) afforded 17 (45 mg, 75%) as

an amorphous white solid. HRMS (ESI): calculated for (M + Na⁺)

C₆₃H₁₀₀N₁₀O₂₈1467.66062, found 1467.65659.

17-Hexa-O-acetate. Per-acetylation of 17, according to the

general procedure, afforded 17-hexa-O-acetate, as an amorphous

white solid in quantitative yield. ¹H NMR (400 MHz, pyridine-d₅,

363 K): δ 8.46 (1H, s, ArH, triazole), 8.37 (2H, m, ArH_XX′), 8.36

(2H, m, ArH_AA′), 7.04 (1H, d, NH_II), 6.9 (1H, br dd, NH_IV), 6.83

(1H, d, J ) 8 Hz, NH_II), 6.69 (2H, m, 2 × NH_I), 6.08 (1H, d, J )

9.8 Hz, NH_IV), 5.95 (1H, br s, H_1I), 5.90 (2H, s, OCH₂), 5.68 (1H,

dd, J ) 9.5, 10.5 Hz, H_3I), 5.64 (1H, d, J ) 2.0 Hz, H_1III), 5.52

(1H, dd, J ) 3, 3 Hz, H_3IV), 5.49 (1H, dd, J ) 9.5, 9.5 Hz, H_4I),

5.46 (1H, dd, J ) 2.0, 5.5 Hz, H_2III), 5.27-5.15 (4H, m, H_4IV′

,

H

_1IV, H_6II, H_5III), 4.94 (1H, dd, J ) 6.5, 14.5 Hz, H_5III′), 4.79 (1H,

dd, J ) 6.6, 5.5 Hz, H_3III), 4.68 (1H, ddd, J ) 3.8, 3.8, 10 Hz,

H_5I), 4.63-4.53 (2H, m, H_4III,H_2I), 4.51 (1H, ddd, J ) 2.0, 6.5,

6.5 Hz, H_5IV), 4.31 (1H, m, H_2IV), 4.09-4.0 (4H, m H_1II, H_3II, H_4II,

H_5II), 3.86 (1H, m, H_6IV), 3.75 (2H, m, H_6I, H_6I′), 3.73 (1H, m,

H_6IV′), 2.45 (3H, s, -OCOCH₃), 2.4 (1H, m, H_2IIeq), 2.35 (3H, s,

-OCOCH₃), 2.17 (3H, s, -OCOCH₃), 2.15 (3H, s, -OCOCH₃), 2.07

(3H, s, -OCOCH₃), 2.05 (3H, s, -OCOCH₃), 2.0 (1H, m, H_2IIax),

1.69 (9H, s, -COC(CH₃)₃), 1.57 (45H, m, 5 × -COC(CH₃)₃). ¹³C

{¹H} NMR (100 MHz, pyridine-d₅, 363 K): δ 171.4-169.0⁴³

(6 × OAc CdO), 165.4⁴³(ipso, Ar) 157.5-156.4⁴³(6 × Boc Cd

O), 151.4⁴³(ipso, Ar) 143.3⁴³(ipso, triazole) 131.5 (ArC_AA′), 126.3

(CH, triazole), 124.5 (ArC_XX′), 108.1 (C_1III), 99.6 (C_1IV), 99.0 (C_1I),

82.9 (C_5II), 80.6 (C_4III), 80.3 (C_4II), 79.9-78.8⁴³(6 × Boc C_q), 77.8

(C_3III), 76.9 (C_6II), 74.9 (C_2III), 74.3 (C_5IV), 73.1 (C_3I), 71.2 (C_4I),

70.5 (C_3IV), 70.4 (C_5I), 67.8 (C_4IV), 59.9 (OCH₂, linker), 54.7 (C_2I),

52.7 (C_5III), 50.7 (C_2IV, C_1II, C_3II), 42.2 (C_6I), 41.5 (C_6IV), 35.8 (C_2II),

29.2-28.7⁴³(6 × Boc (CH₃)₃), 22.2-21.8⁴³(6 × OAc CH₃).

J. Org. Chem, Vol. 72, No. 6, 2007 1977

Quader et al.

(9H, s, -COC(CH₃)₃), 1.61 (9H, s, -COC(CH₃)₃), 1.60 (9H, s, -COC-

(CH₃)₃), 1.58 (9H, s, -COC(CH₃)₃), 1.57 (9H, s, -COC(CH₃)₃), 1.56

(9H, s, -COC(CH₃)₃), 1.10 (3H, s, -CH₃H_19S), 0.91 (3H, s, -CH₃

H_18S). ¹³C {¹H} NMR (100 MHz, pyridine-d₅, 363 K): δ 177.7⁴³

(C_17S), 171.4-169.0⁴³(7 × OAc CdO), 167.2⁴³(CdO, C_21S),

157.5-156.4⁴³(6 × Boc CdO), 144.0⁴³(ipso, triazole) 140.5⁴³

(C_5S), 122.7 (CH, triazole), 109.4 (C_20S) 108.2 (C_1III), 99.8 (C_1IV),

98.7 (C_1I), 82.8 (C_5II), 80.8 (C_4III) ,80.6 (C_4II), 79.9-78.8⁴³(6 ×

Boc C_q), 77.8 (C_3III), 77.0 (C_6II), 74.9 (C_2III), 74.4 (C_5IV, C_3S), 73.3

(C_3I), 70.8 (C_4I), 70.4 (C_3IV, C_5I), 67.9 (C_4IV) ,57.9 (OCH₂, linker),

54.8 (C_2I), 52.4 (C_5III), 50.7 (C_2IV), 50.0 (C_1II, C_3II), 42.4 (C_6I), 41.9

(C_6IV), 38.9 (C_4S), 35.9 (C_2II), 31.3 (C_16S), 29.2-28.7⁴³(6 × Boc

(CH₃)₃), 22.0-20.8⁴³(7 × OAc CH₃) 19.7 (C_19S), 18.8 (C_18S).

HRMS (ESI): calculated for (M + Na⁺) C₉₁H₁₃₉N₉O₃₄1924.93221,

found 1924.92960.

(1H, br dd, NH_I), 6.54 (1H, d, J ) 8.5 Hz, NH_I), 6.44 (1H, d, J )

9 Hz, NH_IV), 6.18 (1H, m, NH_II), 6.05 (1H, s, H_1III), 5.76 (1H, d,

J ) 5.2 Hz, H_2III), 5.66 (1H, dd, J ) 9.4, 10.8 Hz, H_3I), 5.64 (1H,

d, J ) 4 Hz ,H_1I), 5.39 (1H, dd, J ) 9.4, 10 Hz, H_4I), 5.34 (1H, d,

J ) 2.4 Hz, H_1IV), 5.17 (1H, m, H_4IV) 4.95 (1H, dd, J ) 7, 5.3 Hz,

H_3III), 4.71 (2H, m, H_5III, H_5III′), 4.0 (1H,, ddd, J ) 4.4, 4.4, 6.7

Hz, H_4III), 4.55-4.38 (5H, m, H_5II, H_5IV, H_3IV, H_2I, H_5I), 4.30 (1H,

m, H_2IV), 4.14 (2H, m, H_3II, H_4II), 3.84 (1H, m, H_6IV), 3.76-3.60

(3H, m, H_6I, H_6I′, H_6IV′), 3.17 (1H, dd, J ) 6, 4.6 Hz, H_6II), 2.80

(1H, dd, J ) 6, 6 Hz, H_1II), 2.4 (1H, m, H_2IIb), 2.32 (3H, s,

-OCOCH₃), 2.31 (3H, s, -OCOCH₃), 2.16 (3H, s, -OCOCH₃), 2.15

(3H, s, -OCOCH₃), 2.09 (3H, s, -OCOCH₃), 2.12 (1H, m, H_2IIa),

1.66 (9H, s, -COC(CH₃)₃), 1.61 (9H, s, -COC(CH₃)₃), 1.60 (9H, s,

-COC(CH₃)₃), 1.59 (9H, s, -COC(CH₃)₃), 1.58 (9H, s, -COC(CH₃)₃),

1.53 (9H, s, -COC(CH₃)₃). ¹³C {¹H} NMR (100 MHz, pyridine-d₅,

363 K): δ 171.5-170.0⁴³(5 × OAc CdO),162.4⁴³(Boc CdO),

157.4-156.4⁴³(5 × Boc CdO), 104.6 (C_1III), 99.0 (C_1IV,C_1I), 81.34

(C_4III), 79.7-79.0⁴³(6 × Boc C_q), 77.9 (C_3III), 76.9 (C_4II), 76.5

(C_2III), 74.0 (C_5II), 73.2 (C_5IV), 72.9 (C_3I), 71.1 (C_4I), 70.7 (C_5I),

70.2 (C_4IV), 64.9 (C_5III), 60.8 (C_3IV), 54.9 (C_2I), 51.4 (C_2IV), 49.4

(C_3II), 42.2 (C_6I), 41.8 (C_6IV), 38.5 (C_6II), 37.8 (C_1II), 28.6 (C_2II),

29.2-28.6⁴³(6 × Boc (CH₃)₃), 21.7-21.0⁴³(5 × OAc CH₃).

NMR Spectroscopic Data for 22-Tetra-O-acetate: ¹H NMR

(400 MHz, pyridine-d₅, 363 K): δ 6.87 (2H, br m, NH_II,NH_IV),

6.79 (1H, b, J ) 7.8 Hz NH_II), 6.53 (1H,br dd, NH_I), 6.40 (1H, m,

NH_I), 6.21 (1H, d, J ) 9.0 Hz, NH_IV), 5.70 (1H, dd, J ) 9.4, 10.8

Hz, H_3I), 5.57 (1H, d, J ) 3.4 Hz, H_1I), 5.53 (1H, d,, J ) 3.4 Hz,

1,3′′′-Diazido-6-tert-butoxycarbonylamino-3,2′,6′,2′′′,6′′′-penta-

N-(tert-butoxycarbonyl)-1-deamino-6,3′′′-di-deoxyneomycin B

(20), 3′′′-Azido-3,2′,6′,2′′′,6′′′-penta-N-(tert-butoxy-carbonyl)-1,6-

[N-(tert-butoxycarbonyl)epimino]-1-deamino-6,3′′′-di-deoxyneo-

mycin B (21), and 3′,4′,2′′,5′′-Tetra-O-acetyl-1-azido-6-tert-

butoxycarbonylamino-3′′′,4′′′-anhydro-3,6,2′,6′,2′′′,6′′′-penta-N-

(tert-butoxycarbonyl)-1-deamino-6-deoxyneomycin B (22). NaN₃

(11 mg, 0.17 mmol) was added to a solution of 4 (200 mg, 0.17

mmol) in DMF (5 mL) and the resultant mixture was stirred at 50

°C (24 h). The mixture was then diluted with ethyl acetate (50 mL)

and washed with water (2 × 25 mL). The ethyl acetate layer was

dried (Na₂SO₄) and concentrated in vacuo. An aliquot of the crude

reaction (40 mg) was per-acetylated via the general procedure

outlined above and subjected to semipreparative HPLC (MeCN/

H₂O 75:25) to isolate 4-tetra-O-acetate (12 mg), 20-penta-O-acetate

(5 mg) [LRMS (ESI) calculated for (M + Na⁺) C₆₃H₁₀₂N₁₂O₂₈

1497.68, found 1497.94], 21-penta-O-acetate (15 mg) [LRMS (ESI)

calculated for (M + Na⁺) C₆₃H₁₀₁N₉O₂₈1454.67, found 1454.92],

and 22-tetra-O-acetate (3 mg) [LRMS (ESI) calculated for (M +

Na⁺) C₆₁H₉₉N₉O₂₇1412.65, found 1412.7]. Careful column chro-

matography of remaining crude reaction mixture (≈160 mg) on

silica column (DCM/MeOH 98:2 f 95:5) afforded 20 (16 mg,

∼10%) [HRMS (ESI) calculated for (M + Na⁺) C₅₃H₉₂N₁₂O₂₃

1287.62960, found 1287.62327] and 21 (20 mg) [HRMS (ESI)

calculated for (M + Na⁺) C₅₃H₉₁N₉O₂₃1244.61255, found

1244.615243] as white amorphous solids along with some mixed

fraction containing compounds 4, 21, and 22. These fractions were

combined and concentrated and the resulting mixture (120 mg) was

used for the subsequent reactions.

H

_1III), 5.45 (1H, dd, J ) 2.2, 5.5 Hz, H_2III), 5.35 (1H, dd, J ) 9.6,

9.6 Hz, H_4I), 4.71 (1H, d, J ) 3.2 Hz, H_1IV), 4.69-4.57 (3H,m,

H

_3III, H_5III, H_5′III), 4.53-4.35 (8H, m, H_3II, H_4II, H_5II, H_6II, H_2I, H_5I,

_4III, H_2IV), 4.23 (1H, dd, J ) 6, 6 Hz, H_5IV), 4.14 (1H, ddd, J )

4.3, 9.3, 9.4 Hz, H_1II), 3.78 (1H, m, H_6I), 3.76-3.67 (3H, m, H_6IV

,

H_6I′,H_6IV′), 3.50 (1H, dd, J ) 5, 4 Hz, H_3IV), 3.33 (1H, d, J ) 4

Hz, H_4IV), 2.4 (1H, m, H_2IIeq), 2.24 (3H, s, -OCOCH₃), 2.14 (6H,

s, 2 × -OCOCH₃), 2.05 (3H, s, -OCOCH₃), 2.0 (1H, m, H_2IIax),

1.65 (9H, s, -COC(CH₃)₃), 1.64 (9H, s, -COC(CH₃)₃), 1.63 (9H, s,

-COC(CH₃)₃), 1.62 (18H, s, 2 × -COC(CH₃)₃), 1.60 (9H, s, -COC-

(CH₃)₃).

3′,4′,2′′,5′′,4′′′-Penta-O-acetyl-3,2′,6′,2′′′,6′′′-penta-N-(tert-bu-

toxycarbonyl)-1,6-[N-(tert-butoxycarbonyl)epimino]-1-deamino-

6,3′′′-dideoxy3′′′-[4-(4-nitrobenzoyloxymethyl)-[1,2,3]triazol-1-

yl]-neomycin B (23) and 3′,4′,2′′,5′′-Tetra-O-acetyl-3′′′,4′′′-

anhydro-6-tert-butoxycarbonylamino-3,6,2′,6′,2′′′,6′′′-penta-N-

(tert-butoxycarbonyl)-1-deamino-6-deoxy-1-[4-(4-nitrobenzoyl-

oxymethyl)-[1,2,3]triazol-1-yl]-neomycin B (24). A partially

purified mixture of 4, 21, and 22 was subjected to per-acetylation

according to the general procedure. The mixture was then treated

with prop-2-ynyl 4-nitrobenzoate 7 according to the procedure

outlined in the preparation of compound 8. Column chromato-

graphic separation afforded 23-penta-O-acetate (44 mg) [HRMS

(ESI) calculated for (M + H⁺) C₇₃H₁₀₈N₁₀O₃₂1637.72094, found

1637.72108] and 24-tetra-O-acetate (20 mg) [HRMS (ESI) calcu-

lated for (M + Na⁺) C₇₁H₁₀₆N₁₀O₃₁1617.69232, found 1617.69440]

as amorphous white solids.

1

NMR Spectroscopic Data for 20-Penta-O-acetate. H NMR

(400 MHz, pyridine-d₅, 363 K): δ 6.89 (1H, d, J ) 7.8 Hz, NH_II),

6.83 (1H, br dd, NH_IV), 6.77 (1H, d, J ) 8 Hz, NH_I), 6.51 (1H, br

dd, NH_I), 6.44 (2H, m, NH_II, NH_IV), 5.70 (1H, dd, J ) 9.4, 10.7

Hz, H_3I), 5.55 (1H, d, J ) 4 Hz, H_1I), 5.51 (2H, m, H_1III,H_2III),

5.42 (1H, dd, J ) 9.4, 10 Hz, H_4I), 5.15 (2H, m, H_1IV,H_4IV), 4.78

(1H, dd, J ) 6, 5 Hz, H_3III), 4.68 (2H, m, H_5III, H_5III′), 4.56 (1H, m,

H

_4III), 4.52 (2H, m, H_4II,H_5II), 4.48 (1H, m, H_3IV), 4.46 (1H, m,

H_2I), 4.44 (1H, m, H_5I), 4.38 (3H, m, H_3II, H_6II, H_5IV), 4.25 (1H, m,

_2IV), 4.15 (1H, ddd, J ) 3.9, 9.4,9.4 Hz, H_1II), 3.78 (1H, m, H_6IV),

H

1

3.73 (2H, m, H_6I, H_6I′), 3.62 (1H, m, H_6IV′), 2.36 (1H, ddd, dd, J )

4.3,4.3,14 Hz, H_2II), 2.24 (3H, s, -OCOCH₃), 2.23 (3H, s,

-OCOCH₃), 2.14 (3H, s, -OCOCH₃), 2.13 (3H, s, -OCOCH₃), 2.05

(3H, s, -OCOCH₃), 2.12 (1H, m, H_2II′), 1.63 (9H, s, -COC(CH₃)₃),

1.61 (9H, s, -COC(CH₃)₃), 1.59 (18H, s, 2 × -COC(CH₃)₃), 1.56

(18H, s, 2 × -COC(CH₃)₃). ¹³C {¹H} NMR (100 MHz, pyridine-

d₅, 363 K): δ 171.0-169.5⁴³(5 × OAc CdO), 157.0-155.5⁴³(6

× Boc CdO), 108.0 (C_1III), 99.1 (C_1IV, C_1I),81.2 (C_4III), 79.4-78.4⁴³

(6 × Boc C_q), 78.8 (C_4II), 77.2 (C_3III), 76.5 (C_5II), 75.8 (C_2III), 73.2

(C_5IV), 72.5 (C_3I), 70.9 (C_4I), 70.5 (C_5I), 68.2 (C_4IV), 64.6 (C_5III),

60.6 (C_3IV), 57.3 (C_1II) 54.6 (C_2I), 53.6 (C_6II), 51.3 (C_2IV), 50.0 (C_3II),

42.0 (C_6I), 41.3 (C_6IV), 32.0 (C_2II), 28.7-28.3⁴³(6 × Boc (CH₃)₃),

21.0-20.3⁴³(5 × OAc CH₃).

NMR Spectroscopic Data for 23-Penta-O-acetate. H NMR

(400 MHz, pyridine-d₅, 363 K): δ 8.41 (1H, s, ArH, triazole), 8.28

(2H, m, ArH_XX′), 8.22 (2H, m, ArH_AA′), 6.81 (1H, br dd, NH_IV),

6.74 (1H, d, J ) 7 Hz, NH_IV), 6.60 (1H, br dd, NH_I), 6.55 (1H, d,

J ) 8.6 Hz, NH_I), 6.61 (1H, d, J ) 9 Hz, NH_II), 6.02 (1H, br s,

H

_1III), 5.81 (1H, dd, J ) 3, 3 Hz, H_4IV), 5.75 (1H d, J ) 6.2 Hz,

_2III), 5.69 (2H, s, -OCH₂), 5.65 (1H, dd, J ) 9.5, 10.6 Hz, H_3I),

5.65 (2H, m, H_1IV, H_1I), 5.42 (1H, m, H_3IV), 5.37 (1H, dd, J ) 9.5,

10 Hz, H_4I), 4.98 (1H, dd, J ) 3.5, 6.2 Hz, H_3III), 4.96 (1H, m,

H

_5IV), 4.86 (1H, ddd, J ) 2.8, 6.5, 9 Hz, H_2IV), 4.80-4.66 (3H, m,

_5III, H_5′III, H_4III), 4.52 (1H, dd, J ) 4.4, 4.4 Hz, H_5II), 4.48-4.38

(2H, m, H_2I, H_5I), 4.19-4.07 (2H, m, H_3II, H_4II), 3.97 (2H, m, H_6I,

H_6IV), 3.77-3.63 (2H, m, H_6′I, H_6′IV), 3.14 (1H, dd, J ) 6.3, 5 Hz,

H_6II), 2.78 (1H, dd, J ) 5, 5 Hz, H_1II), 2.39 (1H, m, H_2IIb), 2.31

(3H, s, -OCOCH₃), 2.27 (3H, s, -OCOCH₃), 2.15 (6H, m, 2 ×

1

NMR Spectroscopic Data for 21-Penta-O-acetate. H NMR

(400 MHz, pyridine-d₅, 363 K): δ 6.79 (1H,br dd, NH_IV), 6.60

1978 J. Org. Chem., Vol. 72, No. 6, 2007

Multisite Modification of Neomycin B

-OCOCH₃), 2.08 (3H, s, -OCOCH₃), 2.11 (1H, m, H_2IIa), 1.67 (9H,

s, -COC(CH₃)₃), 1.62 (9H, s, -COC(CH₃)₃), 1.58 (9H, s, -COC-

(CH₃)₃), 1.55 (18H, s, 2 × -COC(CH₃)₃), 1.51 (9H, s, -COC(CH₃)₃).

¹³C {¹H} NMR (100 MHz, pyridine-d₅, 363 K): δ 171.0-169.0⁴³

(5 × OAc CdO), 165.0⁴³(Ar_ipso) 162.4⁴³(Boc CdO), 157.5-

(3H, m, -OCOCH₃), 2.14 (3H, s, -OCOCH₃),), 2.06 (3H, s,

-OCOCH₃), 2.0 (1H, m, H_2IIax), 1.66 (9H, s, -COC(CH₃)₃), 1.63

(9H, s, -COC(CH₃)₃), 1.62 (9H, s, -COC(CH₃)₃),1.61 (9H, s, -COC-

(CH₃)₃), 1.55 (9H, s, -COC(CH₃)₃), 1.50 (9H, s, -COC(CH₃)₃); ¹³

C

{¹H} NMR (100 MHz, pyridine-d₅, 363 K): δ 129.6 (ArC_AA′),

123.8 (CH, triazole), 122.2 (ArC_XX′), 108.2 (C_1III), 99.2 (C_1I), 98.9

(C_1IV), 81.3 (C_4III), 79.2 (C_4II), 76.8 (C_3III), 76.5 (C_5II), 76.0 (C_2III),

73.3 (C_5IV), 72.5 (C_3I), 70.8 (C_4I), 70.8 (C_5I), 70.2 (C_4IV), 63.1 (C_5III),

61.4 (C_3IV), 59.5 (OCH₂, linker), 57.2 (C_1II), 54.5 (C_2I), 53.6 (C_6II),

52.5 (C_2IV), 50.2 (C_3II), 42.0 (C_6I, C_6IV), 32.4 (C_2II), 30-27 (6 ×

Boc (CH₃)₃), 21.0-19.0 (5 × OAc CH₃). HRMS (ESI): calculated

for (M + H⁺) C₇₃H₁₀₉N₁₃O₃₂1680.73798, found 1680.74466.

3′′′,4′′′-Anhydro-6-tert-butoxycarbonylamino-3,2′,6′,2′′′,6′′′-

penta-N-(tert-butoxycarbonyl)-6-deoxy-1-N-(propyl)-neomy-

cin B (26). The dianhydro neomycin 4 (50 mg 0.04 mmol) was

stirred at room temperature in neat propylamine (1 mL) for 16 h.

The mixture was then evaporated to dryness and column chro-

matographic separation (DCM/MeOH 97:5 f 95:8) afforded 26

(25 mg, 47%) as a white amorphous solid. LRMS (ESI): calculated

for (M + H⁺) C₅₆H₉₉N₇O₂₃1238.69, found 1239.0.

156.2⁴³(5 × Boc CdO), 151.35⁴³(Ar_ipso) 143.13⁴³(triazole_ipso

)

131.3 (ArC_AA′), 125.58 (CH, triazole), 124.0 (ArC_XX′), 104.7 (C_1III),

99.1 (C_1IV, C_1I), 81.9 (C_4III), 79.9-78.9⁴³(6 × Boc C_q), 77.5 (C_3III),-

76.6 (C_4II), 76.34 (C_2III), 73.57 (C_5II), 73.56 (C_5IV), 72.7 (C_3I), 71.2

(C_4I), 70.8 (C_5I), 70.3 (C_4IV), 65.0 (C_5III), 61.5 (C_3IV), 59.57 (OCH₂,

linker), 54.6 (C_2I), 52.7 (C_2IV) 49.2 (C_3II), 42.1 (C_6I), 41.9 (C_6IV),

38.1 (C_6II), 37.88 (C_1II), 28.0 (C_2II), 29.3-28.7⁴³(6 × Boc (CH₃)₃),

22.0-21.0⁴³(5 × OAc CH₃).

1

NMR Spectroscopic Data for 24-Tetra-O-acetate. H NMR

(400 MHz, pyridine-d₅, 363 K): δ 8.31 (2H, m, ArH_XX′), 8.26 (2H,

m, ArH_AA′), 8.06 (1H, s, ArH, triazole), 6.95 (1H, m, NH_II), 6.87

(3H, br m, NH_I), 6.80 (1H, m, NH_I), 6.63 (2H, m, NH_II,NH_IV),

6.21 (1H, d, J ) 9.4 Hz, NH_IV), 5.63 (1H dd, J ) 9.5, 10.8 Hz,

H_3I), 5.66 (2H, s,OCH₂), 5.60 (1H, d, J ) 3.8, Hz, H_1I), 5.55 (1H,

d, J ) 2.3 Hz, H_1III), 5.47 (1H, ddd, J ) 3.9, 10.4, 10.4 Hz, H_1II),

5.44 (1H, dd,, J ) 2.4, 5.3 Hz, H_2III), 5.42 (1H, dd, J ) 9.5, 9.5

Hz, H_4I), 4.91 (1H, ddd, J ) 2.9, 9.2, 11.6 Hz, H_6II), 4.68-4.60

(7H, m, H_1IV,H_3IIIH_5IIIH_5′IIIH_3IIH_4IIH_5II), 4.54-4.46 (3H, m,

N-Acetyl-26-tetra-O-acetate. Per-acetylation of compound 26

according to the general method afforded N-acetyl-26-tetra-O-

1

acetate. H NMR (400 MHz, pyridine-d₅, 373 K): δ 6.77 (1H, br

H_4III, H_2I, H_5I), 4.39 (1H, ddd,, J ) 9, 4.9, 3.6, Hz, H_2IV), 4.22

dd, NH_IV), 6.53 (1H, br dd, NH_I), 6.44 (2H,m, NH_I, NH_II), 6.25

(1H, m, NH_II), 6.10 (1H, J ) 9.5 Hz, NH_IV), 5.72 (1H, dd, J )

9.4, 10.9 Hz, H_3I), 5.58 (1H, dd, J ) 2, 5 Hz, H_2III), 5.53 (1H, d,

J ) 4 Hz, H_1I), 5.48 (1H, br s, H_1III), 5.38 (1H, dd, J ) 9.5, 910

Hz, H_4I), 4.81 (1H, m, H_3III), 4.79 (1H, d, J ) 3.3 Hz, H_1IV), 4.71-

(1H, dd,, J ) 6, 6 Hz, H_5IV), 3.81 (1H, m, H _6I), 3.75-3.71 (3H,

m, H_6IV,H_6I’H_6IV′), 3.49 (1H, dd, J ) 5.2, 3.9 Hz, H_3IV), 3.33 (1H,

d, J ) 3.8 Hz, H_4IV), 2.82 (1H, ddd, J ) 13.7, 10.4, 4.3 Hz, H_2IIax),

2.65 (1H, ddd, J ) 3.8, 3.8, 13.7 Hz, H_2IIeq), 2.32 (3H, s,

-OCOCH₃), 2.29 (3H, s, -OCOCH₃), 2.12 (3H, m, -OCOCH₃), 2.03

(3H, s, -OCOCH₃), 1.64 (9H, s, -COC(CH₃)₃), 1.62 (18H, s, 2 ×

-COC(CH₃)₃), 1.69 (27H, m, 3 × -COC(CH₃)₃). ¹³C {¹H} NMR

(100 MHz, pyridine-d₅, 363 K): δ 171.0-170.0⁴³(4 × OAc Cd

4.68 (4H,br m, H_1II, H_3II, H_5III, H_5′III), 4.61-4.49 (3H, br m, H_4III

,

H_6II, H_2IV), 4.49-4.39 (4H, br m, H_5II, H_4II, H_2I, H_5I), 4.32 (1H,

dd,, J ) 6, 6 Hz, H_5IV), 3.77 (4H, m, H_6I, H_6IV, H_6I′, H_6IV′), 3.55

(1H, dd, J ) 5, 4 Hz, H_3IV), 3.37 (1H, d, J ) 4 Hz, H_4IV), 3.34

(2H, m, -NAc-CH₂-), 2.57 (1H, m, H_2IIeq), 2.37 (3H, s, -OCOCH₃),

2.36 (1H, m, H_2IIax), 2.33 (2H,br s, -CH₂-), 2.18 (6H, s,2 ×

-OCOCH₃), 2.15 (3H, s, -OCOCH₃), 2.07 (3H, s, -OCOCH₃), 1.67

(9H, s, -COC(CH₃)₃), 1.64 (9H, s, -COC(CH₃)₃), 1.64 (2H, s, (-NAc-

CH₂-CH₂-),1.63 (27H, s, 3 × -COC(CH₃)₃), 1.60 (9H, s, -COC-

(CH₃)₃), 1.0 (-CH₃); ¹³C {¹H} NMR (100 MHz, pyridine-d₅, 363

K): δ 173.66⁴³(NAc, CdO),171.21-170.0⁴³(4 × OAc CdO),

157.2-155.3⁴³(6 × Boc CdO), 108.4 (C_1III), 99.9 (C_1IV), 98.9 (C_1I),

81.0 (C_4III), 79.6 (C_4II), 76.1 (C_5II), 79.7-78.0⁴³(6 × Boc C_q), 76.0

(C_3III), 75.4 (C_2III), 72.9 (C_5IV), 72.3 (C_3I), 71.1 (C_5I), 70.9 (C_4I),

70.2 (C_5I), 64.2 (C_5III), 54.7 (C_2I), 52.7 (C_4IV), 52.2 (C_3IV), 51.8

(C_6I!), 50.8 (C_1II, C_3II), 47.5 (-NAc-CH₂-), 46.3 (C_2IV), 43.7 (C_6IV),

42.1 (C_6I), 31.5 (C_2II), 29.2-27.2⁴³(6 × Boc (CH₃)₃), 22.0-20.5⁴³

(5 × Ac, CH₃), 24.2 (-NAc-CH₂-CH₂-), 11.4 (-CH₃). HRMS

(ESI): calculated for (M + Na⁺) C₆₆H₁₀₉N₇O₂₈1470.72183, found

1470.72365.

O), 165.0⁴³(Ar_ipso) 157.4-156.4⁴³(6 × Boc CdO), 151.4⁴³(Ar_ipso

)

143.1⁴³(triazole_ipso) 131.6 (ArC_AA′), 123.9 (ArC_XX′), 123.3 (CH,

triazole), 108.7 (C_1III), 99.7 (C_1IV) 99.0 (C_1I), 81.1 (C_4III), 80.1 (C_5II),

79.9-78.9⁴³(6 × Boc C_q), 76.5 (C_4II,C_3III), 75.8 (C_2III), 72.5 (C_5IV),

72.2 (C_3I), 70.8 (C_4I, C_5I), 64.2 (C_5III), 59.9 (OCH₂, linker), 56.9

(C_1II), 54.9 (C_2I), 53.2 (C_6II), 52.6 (C_4IV), 51.9 (C_3IV), 50.6 (C_3II),

46.3 (C_2IV), 42.9 (C_6IV), 41.9 (C_6I), 34.3 (C_2II), 29.14-28.6⁴³(6 ×

Boc (CH₃)₃), 21.5-21.1⁴³(4 × OAc CH₃).

3′,4′,2′′,5′′,4′′′-Penta-O-acetyl-1-azido-6-tert-butoxycarbonyl-

amino-3,2′,6′,2′′′,6′′′-penta-N-(tert-butoxycarbonyl)-1-deamino-

6,3′′′-dideoxy-3′′′-[4-(4-nitrobenzoyloxymethyl)-[1,2,3]-triazol-

1-yl]-neomycin B (25). NaN₃(3 mg) was added to a solution of

23-penta-O-acetate (20 mg, 0.17 mmol) in DMF (1 mL) and the

mixture was stirred at 50 °C (12 h). The reaction mixture was

diluted with ethyl acetate (10 mL) and washed with water (2 × 10

mL). The ethyl acetate layer was dried (Na₂SO₄) and concentrated

in vacuo. Flash column chromatography afforded 25-penta-O-

acetate (20 mg, 98%) as an amorphous white solid. ¹H NMR (400

MHz, pyridine-d₅, 363 K): δ 8.40 (1H, s, ArH, triazole), 8.28 (2H,

m, ArH_XX′), 8.22 (2H, m, ArH_AA′), 6.84 (1H, d, J ) 7 Hz, NH_IV),

6.74 (3H, m, NH_IV,2XNH_II), 6.61 (1H, br dd, NH_I), 6.46 (1H, d,

J ) 8.0 Hz, NH_I), 5.76 (1H, dd, J ) 3, 3 Hz, H_4IV), 5.71 (1H dd,

J ) 9.2, 10.8 Hz, H_3I), 5.69 (2H, s,OCH₂), 5.58 (1H, d, J ) 2.8,

Hz, H_1I), 5.54 (1H, d, J ) 3.7 Hz, H_1IV), 5.51 (2H, m, H_1III, H_2III),

5.44 (1H, m, H_3IV), 5.41 (1H, dd, J ) 9.4, 10 Hz, H_4I), 4.92 (1H,

ddd, J ) 2.6, 6.6, 6.6 Hz, H_5IV), 4.86 (1H, m, H_2IV), 4.84 (1H, m,

Acknowledgment. We gratefully acknowledge financial

support for this work from Eskitis Institute, Natural Product

Discovery, and Griffith University. We also gratefully acknowl-

edge the Eskitis Institute and Griffith University for the generous

provision of postgraduate student scholarships (S.Q.).

Supporting Information Available: General experimental

procedures, 1D and 2D NMR spectra of the per-acetylated

aminoglycosides 2, 4, 5, 8, 13, 14, and 16-26 and the starting

synthons 9 and 10. This material is available free of charge via the

Internet at http://pubs.acs.org.

H_3III), 4.73 (1H, m, H_5III), 4.63 (2H, m, H_5′III, H_4III), 4.52-4.34 (6H,

m, H_2I, H_3II, H_4II, H_5II, H_6II, H_5I), 4.46 (1H, ddd, J ) 4, 9.6, 9.6 Hz,

H_1II), 3.93 (1H, m, H_6IV), 3.71 (3H, m, H_6I, H_6I′, H_6IV′), 2.35 (1H,

m, H_2IIeq), 2.26 (3H, s, -OCOCH₃), 2.21 (3H, s, -OCOCH₃), 2.16

JO0620967

J. Org. Chem, Vol. 72, No. 6, 2007 1979

Article Doi

Multisite modification of neomycin B: Combined Mitsunobu and click chemistry approach

DOI: 10.1021/jo0620967

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