Azidomethyl 4-Nitrophenyl Carbonate - A Reagent for the One-Step Introduction of the Azidomethyloxycarbonyl (Azoc) Protecting Group

Abstract

Presented here is the three-step synthesis of azidomethyl 4-nitrophenyl carbonate in 58% overall yield. This carbonate allows for the high-yielding (³90%) introduction of the phosphine-labile azidomethyloxycarbonyl (Azoc) protecting group in one step. The reagent protects a range of amines, including amino acids. For nonionic substrates, pure carbamates are obtained after extractive workup.

Protecting groups are indispensable tools for the synthesis of complex target molecules. Ideal protecting groups should be (i) easy to introduce in high yield, (ii) stable toward a wide range of reaction conditions, (iii) easily removed in high yield, and (iv) orthogonal to as many other protecting groups as possible. [¹] The ever-increasing number of protecting groups makes the latter criterion increasingly difficult to meet. Recently, Pothukanuri and Winssinger introduced the azidomethyloxycarbonyl (Azoc) group as a new protecting group for amines and alcohols. [²] This group can be removed by treatment with phosphines. It has been succesfully employed for the synthesis of peptides, carbohydrates, peptide nucleic acids (PNAs), and triazoles. [²-4] The Azoc group satisfactorily fulfills criteria (ii)-(iv), but thus far, the introduction of Azoc moieties required a two-step process (Scheme  [¹] ) [²] [³] with overall yields of 34-86% that often involves two chromatographic purifications.

Here we present the synthesis of azidomethyl 4-nitrophenyl carbonate (1), a reagent for the one-step introduction of the Azoc group, together with a convenient methodology for protecting amines using this reagent (Table  [¹] ).

The choice of the leaving group affects the stability of a given Azoc reagent. To be nonexplosive, organic azides should fulfill the criteria that the number of nitrogen atoms must not exceed that of carbons and that (N _C + N _O )/N _N ÷ 3, [5] where N _C , N _O , and N _N are the number of carbon atoms, oxygen atoms, and nitrogen atoms, respectively. Based on this equation, leaving groups like HOAt [(N _C + N _O )/N _N  = 1.4], HOBt [(N _C + N _O )/N _N  = 1.8], and NHS [(N _C + N _O )/N _N  = 2.8] were excluded. Instead, 4-nitrophenol [(N _C + N _O )/N _N  = 3.3] and pentafluorophenol [(N _C + N _O + N _F )/N _N  = 5.3] were identified as leaving-group options. A second criterion helped to select the former among these two options. This was the desire to suppress the formation of potentially unstable, toxic and explosive side products like azidomethyl azidoformate [(N _C + N _O )/N _N  = 0.7] and carbonyl diazide (Scheme  [²] ).

The pK _a values as a simple measure of leaving group capability are 7.16 for 4-nitrophenol and 5.30 for pentafluorophenol, making carbonates of 4-nitrophenol the more stable of the two, less likely to undergo exchange reactions producing azidocarbonates. Additionally, formation of chloromethyl 4-nitrophenyl carbonate (3), iodomethyl 4-nitrophenyl carbonate (4), and substitution reactions with iodo compound 4 were described in the literature, particularly the patent literature, making a successful synthesis of 1 likely. [6]

Table 1 Azoc Protection According to General Procedure [¹¹] ^a

Compd	Product	Temp (˚C)	Time	Yield (%)
6a		0	5 min	>99
6b		0	5 min	>99
6c		20	24 h	b
6d		0	5 min	98
6e		20	24 h	c
6f		0	20 min	93
6g d		20	1 h	90e
6h		20	30 min	99f
6i		20	5 min	91g
a X = NH, O b No conversion observed after 24 h at 20 ˚C or after 6 h at 60 ˚C. c No conversion observed after 24 h. d Starting material 5g (hydrochloride) was synthesized according to ref. 12. e Yield after flash chromatography, see footnote 11 for details. f Different protocol, see footnote 13. g Different procedure, see footnote 14.

Preparation of 1 (Scheme  [³] ) started from commercially available chloromethyl chloroformate (2), which gave nitrophenolate 3 in 89% yield. [6b] Treatment of 3 with sodium azide in DMF, acetonitrile, or toluene/15-crown-5, respectively, showed products of addition-elimination reactions at the carbonyl group and decomposition of 3, without formation of the desired product 1. But iodide 4 was accessible in 90% yield through a Finkelstein reaction. [6b] Despite its better leaving group, compound 4 reacted similarly to chloride 3 when treated with sodium azide. Only when the iodide leaving group of 4 was activated by addition of Ag⁺ ions did the methylene carbon reach the level of reactivity required to compete successfully with that of the carbonate carbon.

Literature reports exist on the substitution of the iodine in 4 in the presence of AgCO₃ as base and with silver salts of nucleophiles. [6b-6d] We tested silver azide [7] [8] and conditions described by Gediya et al., [6c] as well as those described by Gallop et al. [6d] We observed formation of the desired azidomethyl 4-nitrophenyl carbonate 1 in 20% [9] yield under the former conditions that involve acetone as solvent, and in 72% yield under the latter conditions that involve toluene as solvent. [¹0]

With azidomethyl 4-nitrophenyl carbonate (1) in hand, we then optimized the conditions for the protection of amines. With diisopropyl ethyl amine as base and DMF as solvent, quantitative conversions were observed. Diluting the reaction mixture with ethyl acetate, followed by washing with aqueous carbonate, water, and brine gave carbamates of amines with nonionic residues in near-quantitative yield (Table  [¹] ). [¹¹]

Neither anilinic amine (5b), aniline itself (5c), nor benzyl alcohol (5e) showed any reactivity under the chosen conditions, allowing for chemoselective protection of amino groups in richly functionalized substrates. The more electron-rich aminopyrrole 5g [¹²] was successfully protected, though. Also, the protection of ionic substrates, such as phosphate 5h [¹³] or β-alanine (5i) [¹4] was successful, and pure products were obtained after elution from ion-­exchange columns. The one-step protocol avoids the de­esterification step (88-95% yield) of the known syntheses. [²]

In conclusion, we report the three step syntheses of azidomethyl 4-nitrophenyl carbonate (1) in 58% overall yield. Compound 1 was employed for chemoselective one-step Azoc protection of amines in yields of 90-99%, including protection of polar amines like phosphate 5h and amino acid 5i. It is hoped that these results will lead to a more widespread use of the Azoc protecting group. Phosphine-labile protecting groups and linkers are attractive for their orthogonality to other functionalities and their compatibility with complex substrates, including nucleic acids. [¹5]

Supporting Information for this article is available online at http://www.thieme-connect.com.accesdistant.sorbonne-universite.fr/ejournals/toc/synlett. Included are spectroscopic data of compounds 1, 6b,d,g-i, and the complete author list of reference 15.

Acknowledgment

This work was supported by DFG (Grants No. RI 1063/8-1 and RI 1063/9-1 to C.R.). The authors thank K. Imhof for providing the precursor of compound 5g and E. Kervio, H. Griesser, and R. Haug for helpful discussions.

References and Notes

• 1 Isidro-Llobet A. Álvarez M. Albericio F. Chem. Rev.  2009,  109:  2455 
  CrossrefPubMedSearch in Google Scholar
• 2 Pothukanuri S. Winssinger N. Org. Lett.  2007,  9:  2223 
  CrossrefSearch in Google Scholar
• 3 Pothukanuri S. Pianowski Z. Winssinger N. Eur. J. Org. Chem.  2008,  3141 
  Crossref
• 4 Loren JC. Krasinski A. Fokin VV. Sharpless KB. Synlett  2005,  2847 
  Thieme Connect
• 5a Smith PAS. Open-Chain Nitrogen Compounds   Vol. 2:  Benjamin; New York: 1966.  p.211-256  
  Search in Google Scholar
• 5b Bräse, S.; Gil, C.; Knepper, K.; Zimmermann, V. Angew. Chem. Int. Ed. 2005, 44, 5188
• 5c Bräse S. Bannert K. Organic Azides: Syntheses and Applications   Vol. 1:  Wiley; Chichester: 2010.  p.3-27  
  Search in Google Scholar
• See, for example:
• 6a Gangwar S. Pauletti GM. Siahaan TJ. Stella VJ. Borchardt RT. J. Org. Chem.  1997,  62:  1356 
  Search in Google Scholar
• 6b Goebel T, Humbert-Droz E, and Schwarzenbach M. inventors; WO  200029378.  ; Chem. Abstr. 2000, 132, 347496
• 6c Gediya LK. Khandelwal A. Patel J. Belosay A. Sabnis G. Mehta J. Purushottamachar P. Njar VCO. J. Med. Chem.  2008,  51:  3895 
  Search in Google Scholar
• 6d Gallop MA, Cundy KC, Zhou CX, Yao F, Xiang J.-N, Ollman IR, and Qiu FG. inventors; US 20060229361  A1.  ; Chem. Abstr. 2006, 145, 418708
• 12 Baird E. Dervan PB. J. Am. Chem. Soc.  1996,  118:  6141 
  CrossrefSearch in Google Scholar
• 15 Bentley DR. et al. Nature (London)  2008,  456:  53 
  CrossrefSearch in Google Scholar

CAUTION: Silver azide is an explosive. Safety equipment such as leather gloves, face shield, protective shield, and ear plugs is recommended. Further safety notes for work with metal and organic azides can be found in ref. 5c.

Remaining silver azide may be quenched through oxidation with aq KI₃ solution under sulfide catalysis (iodine-azide reaction): Silver azide containing salts and contaminated filters were added to a stirred aq solution of KI₃ (500 mL, 0.2 M) and Na₂S (50 mg) resulting in immediate development of N₂. This slurry was stirred overnight.

Calculated from signals in ^¹H NMR spectrum of crude.

Azidomethyl 4-Nitrophenyl Carbonate (1)
AgNO₃ (4.60 g, 27.1 mmol, 1.25 equiv) was dissolved in H₂O (20 mL) and added to a stirred solution of NaN₃ (1.76 g, 27.1 mmol, 1.25 equiv) in H₂O (20 mL). AgN₃ (4.06 g, 27.1 mmol, 1.25 equiv) immediately formed as a white microcrystalline precipitate. The supernatant was aspired with a syringe, and the precipitate was washed with H₂O (3 × 30 mL), acetone (3 × 30 mL), and toluene (3 × 30 mL). The AgN₃ was suspended in toluene (50 mL), MS 4 Å (6.0 g) were added, and the resulting slurry was stirred 15 min under argon. Iodomethyl 4-nitrophenyl carbonate^6b (4, 7.00 g, 21.67 mmol, 1.0 equiv) was added in one portion. Complete conversion was detected by TLC [R _f = 0.52 (CH₂Cl₂-PE = 3:2)] after 3 h of stirring in the dark. Silver salts (compare ref. 8) were filtered off, and the filtrate was diluted with CH₂Cl₂ (200 mL) and washed with H₂O (50 mL), Na₂CO₃ solution (2 × 50 mL, 2 M), H₂O (50 mL), and sat. NaCl solution (50 mL). The organic phase was dried over Na₂SO₄, filtered, and evaporated under reduced pressure. The resulting yellow oil was purified by flash column chromatography, eluting with CH₂Cl₂-PE (3:4) → CH₂Cl₂-PE (1:1). The title compound was obtained as a slight yellowish oil that slowly crystallized (3 d) at -18 ˚C, but rapidly crystallized after addition of a seed crystal (3.72 g, 15.6 mmol, 72%). Yellowish powder, mp 35 ˚C. IR (neat): 3119, 2962, 2559, 2369, 2105, 1967, 1766, 1617, 1594, 1523, 1491, 1347, 1198 cm^-¹. ^¹H NMR (300 MHz, CDCl₃): δ = 5.32 (s, 2 H, CH₂N₃), 7.40-7.46 (m, 2 H, ArH), 8.28-8.34 (m, 2 H, ArH). ^¹³C NMR (75 MHz, CDCl₃): δ = 78.96 (CH₂), 121.74 (CH), 125.45 (CH), 145.72, 152.29, 155.04. MS (EI, 70 eV): m/z (%) = 238 (2) [M]⁺, 196 (12), 166 (36), 139 (40), 122 (42), 56 (100), 28 (71). HRMS (EI): m/z calcd for C₈H₆N₄O₅: 238.0338 [M]⁺; found: 238.0319 [M]⁺. Anal. Calcd for C₈H₆N₄O₅: C, 40.35; H, 2.54; N, 23.53. Found: C, 40.25; H, 2.69; N, 23.26.

General Procedure for Azoc Protection with Azidomethyl 4-Nitrophenyl Carbonate (1) and Analytical Data of Compounds 6b,d,g Azidomethyl 4-nitrophenyl carbonate (1, 300 mg, 1.26 mmol, 1.0 equiv) was dissolved in anhyd DMF (3 mL) and cooled to 0 ˚C. Either of compounds 5a-g (1.26 mmol) was dissolved in anhyd DMF (2 mL) in an separate flask and treated with DIPEA (1.39 mmol, 236 µL, 1.1 equiv). The resulting solution was added dropwise, within 2 min, to the solution of 1. The reaction was monitored via TLC (CH₂Cl₂-PE = 3:2). After complete conversion, the reaction mixture was diluted with EtOAc (200 mL) and washed with Na₂CO₃ solution (8 × 50 mL, 2 M). The combined aqueous solutions were re-extracted with EtOAc (30 mL). The extract was washed once with H₂O (10 mL). The combined organic phases were washed with sat. NaCl solution (2 × 50 mL), dried over Na₂SO₄, filtered, concentrated in vacuo and dried at <10^-³ mbar for at least 3 h. Analytical data of compounds 6a and 6f were found to be identical to those in ref. 2.
Azidomethyl 4-Aminobenzylcarbamate (6b) Orange oil which slowly crystallized at -18 ˚C (2 weeks) to give an orange solid with mp 26-28 ˚C. IR (neat): 3423, 3328, 2154, 2098, 1723, 1697, 1617, 1539, 1512 cm^-¹. ^¹H NMR (300 MHz, CDCl₃): δ = 3.68 (s, 2 H, NH₂), 4.27 (d, ^³ J = 5.9 Hz, 2 H, CH₂Ar), 5.05-5.20 (m, 3 H, CH₂N₃, NH), 6.62-6.68 (m, 2 H, ArH), 7.08 (d, ^³ J = 8.3 Hz, 2 H, ArH). ^¹³C NMR (75 MHz, CDCl₃): δ = 44.89 (ArCH₂), 75.32 (CH₂N₃), 115.25, 127.59, 128.72, 129.04, 146.08, 155.18. ESI-HRMS: m/z calcd for C₉H₁₁N₅O₂Na: 244.0805 [M + Na]⁺; found: 244.0802 [M + Na]⁺.
Azidomethyl 5-Hydroxypentylcarbamate (6d) Colorless oil. IR (neat): 3329, 2937, 2864, 2140, 2100, 1705, 1532 cm^-¹. ^¹H NMR (300 MHz, CDCl₃): δ = 1.36-1.48 (m, 2 H, H-3), 1.51-1.65 (m, 5 H, H-2, H-4, OH), 3.24 (q, ^³ J = 6.7 Hz, 2 H, CH₂NH), 3.65 (t, ^³ J = 6.4 Hz, 2 H, CH₂OH), 5.02 (s, 1 H, NH), 5.13 (s, 2 H, CH₂N₃). ^¹³C NMR (75 MHz, CDCl₃): δ = 22.87 (C-3), 29.51, 32.13, 41.02 (C-1), 62.55 (C-5), 75.22 (CH₂N₃), 155.36 (C=O). ESI-HRMS: m/z calcd for C₇H₁₄N₄O₃Na: 225.0958 [M + Na]⁺; found: 225.0964 [M + Na]⁺.
Methyl 4-[(Azidomethoxy)carbonylamino]- N -methyl-pyrrole-2-carboxylate (6g) Compound 6g was purified via flash column chromatography using a gradient of CH₂Cl₂ → CH₂Cl₂-MeOH (99:1).
Orange brownish powder; mp 136-138 ˚C. IR (neat): 3509, 3310, 2958, 2163, 2141, 2112, 1967, 1717, 1679, 1586, 1565 cm^-¹. ^¹H NMR (500 MHz, DMSO-d ₆): δ = 3.72 (s, 3 H, Me), 3.82 (s, 3 H, Me), 5.27 (s, 2 H, CH₂N₃), 6.69 (d, ⁴ J = 1.9 Hz, 1 H, ArH), 7.14 (d, ⁴ J = 1.9 Hz, 1 H, ArH), 9.80 (s, 1 H, NH). ^¹³C NMR (126 MHz, DMSO-d ₆): δ = 36.09, 50.88, 74.88, 107.56, 118.99, 119.47, 121.92, 152.38, 160.54. ESI-HRMS: m/z calcd for C₉H₁₁N₅O₄Na: 276.0703 [M + Na]⁺; found: 276.0716 [M + Na]⁺.

Ammonium 2-[(Azidomethoxy)carbonylamino]ethyl Hydrogenphosphate (6h) Azidomethyl 4-nitrophenyl carbonate (1, 300 mg, 1.26 mmol, 1.0 equiv) and 2-aminoethyl dihydrogen phosphate (177.7 mg, 1.26 mmol, 1.0 equiv) were added to a flask and suspended in DMF (4 mL). DIPEA (4.16 mmol, 707 µL, 3.3 equiv) and H₂O (3 mL) were added. The resulted suspension was sonicated for 2 min and then stirred at 20 ˚C. After 20 min, a clear solution formed. A complete conversion was detected after 30 min via TLC (CH₂Cl₂-PE = 3:2). The reaction mixture was diluted with H₂O (200 mL) and poured on a DEAE-cellulose column [100 g, 20 cm, HCO₃ ^- form, Express-Ion exchanger D, Whatman (Maidstone, UK)]. The column was washed with H₂O (200 mL) and 100 mM NH₄HCO₃ solution (500 mL). The product-containing fractions were combined and the resulting solution concentrated to 20 mL, followed by lyophilization to dryness. The title compound 6h was obtained as a colorless solid (321 mg, 1.25 mmol, 99%). IR (neat): 3210, 2889, 2103, 1705, 1532, 1455 cm^-¹. ^¹H NMR (300 MHz, D₂O): δ = 3.28 (t, ^³ J = 5.3 Hz, 2 H), 3.75-3.83 (m, 2 H), 5.06 (s, 2 H, CH₂N₃). ^¹³C NMR (75 MHz, D₂O): δ = 41.25 (d,
^³ J _P-C2 = 7.8 Hz, C-2), 63.59 (d, ^² J _P-C1 = 5.2 Hz, C-1), 75.71 (CH₂N₃), 157.35 (C=O). ^³¹P NMR (121.5 MHz, D₂O): δ = 1.10. ESI-HRMS: m/z calcd for C₄H₈N₄O₆P: 239.0187 [M]^-; found: 239.0175 [M]^-.

Ammonium 3-[(Azidomethoxy)carbonylamino]-propanoate (6i)
A sample of β-alanine (112.2 mg, 1.26 mmol) was treated as described in ref. 13 but in the presence of a smaller amount of DIPEA (471 µL, 2.77 mmol, 2.2 equiv). The column purification used a higher concentration of NH₄HCO₃ (250 mM) for elution from the DEAE-cellulose. The title compound was obtained as colorless oil (236.2 mg, 1.15 mmol, 91%). IR (neat): 3231, 2965, 2103, 1967, 1701, 1524, 1447, 1409, 1219 cm^-¹. ^¹H NMR (300 MHz, D₂O): δ = 2.44 (t, ^³ J = 6.6 Hz, 2 H), 3.31 (t, ^³ J = 6.6 Hz, 2 H), 5.06 (s, 2 H, CH₂N₃). ^¹³C NMR (75 MHz, D₂O): δ = 34.95, 36.79, 75.62 (CH₂N₃), 157.14, 177.60. ESI-HRMS: m/z calcd for C₅H₇N₄O₄: 187.0473 [M]^-; found: 187.0468 [M]^-.

References and Notes

• 1 Isidro-Llobet A. Álvarez M. Albericio F. Chem. Rev.  2009,  109:  2455 
  CrossrefPubMedSearch in Google Scholar
• 2 Pothukanuri S. Winssinger N. Org. Lett.  2007,  9:  2223 
  CrossrefSearch in Google Scholar
• 3 Pothukanuri S. Pianowski Z. Winssinger N. Eur. J. Org. Chem.  2008,  3141 
  Crossref
• 4 Loren JC. Krasinski A. Fokin VV. Sharpless KB. Synlett  2005,  2847 
  Thieme Connect
• 5a Smith PAS. Open-Chain Nitrogen Compounds   Vol. 2:  Benjamin; New York: 1966.  p.211-256  
  Search in Google Scholar
• 5b Bräse, S.; Gil, C.; Knepper, K.; Zimmermann, V. Angew. Chem. Int. Ed. 2005, 44, 5188
• 5c Bräse S. Bannert K. Organic Azides: Syntheses and Applications   Vol. 1:  Wiley; Chichester: 2010.  p.3-27  
  Search in Google Scholar
• See, for example:
• 6a Gangwar S. Pauletti GM. Siahaan TJ. Stella VJ. Borchardt RT. J. Org. Chem.  1997,  62:  1356 
  Search in Google Scholar
• 6b Goebel T, Humbert-Droz E, and Schwarzenbach M. inventors; WO  200029378.  ; Chem. Abstr. 2000, 132, 347496
• 6c Gediya LK. Khandelwal A. Patel J. Belosay A. Sabnis G. Mehta J. Purushottamachar P. Njar VCO. J. Med. Chem.  2008,  51:  3895 
  Search in Google Scholar
• 6d Gallop MA, Cundy KC, Zhou CX, Yao F, Xiang J.-N, Ollman IR, and Qiu FG. inventors; US 20060229361  A1.  ; Chem. Abstr. 2006, 145, 418708
• 12 Baird E. Dervan PB. J. Am. Chem. Soc.  1996,  118:  6141 
  CrossrefSearch in Google Scholar
• 15 Bentley DR. et al. Nature (London)  2008,  456:  53 
  CrossrefSearch in Google Scholar

CAUTION: Silver azide is an explosive. Safety equipment such as leather gloves, face shield, protective shield, and ear plugs is recommended. Further safety notes for work with metal and organic azides can be found in ref. 5c.

Remaining silver azide may be quenched through oxidation with aq KI₃ solution under sulfide catalysis (iodine-azide reaction): Silver azide containing salts and contaminated filters were added to a stirred aq solution of KI₃ (500 mL, 0.2 M) and Na₂S (50 mg) resulting in immediate development of N₂. This slurry was stirred overnight.

Calculated from signals in ^¹H NMR spectrum of crude.

Azidomethyl 4-Nitrophenyl Carbonate (1)
AgNO₃ (4.60 g, 27.1 mmol, 1.25 equiv) was dissolved in H₂O (20 mL) and added to a stirred solution of NaN₃ (1.76 g, 27.1 mmol, 1.25 equiv) in H₂O (20 mL). AgN₃ (4.06 g, 27.1 mmol, 1.25 equiv) immediately formed as a white microcrystalline precipitate. The supernatant was aspired with a syringe, and the precipitate was washed with H₂O (3 × 30 mL), acetone (3 × 30 mL), and toluene (3 × 30 mL). The AgN₃ was suspended in toluene (50 mL), MS 4 Å (6.0 g) were added, and the resulting slurry was stirred 15 min under argon. Iodomethyl 4-nitrophenyl carbonate^6b (4, 7.00 g, 21.67 mmol, 1.0 equiv) was added in one portion. Complete conversion was detected by TLC [R _f = 0.52 (CH₂Cl₂-PE = 3:2)] after 3 h of stirring in the dark. Silver salts (compare ref. 8) were filtered off, and the filtrate was diluted with CH₂Cl₂ (200 mL) and washed with H₂O (50 mL), Na₂CO₃ solution (2 × 50 mL, 2 M), H₂O (50 mL), and sat. NaCl solution (50 mL). The organic phase was dried over Na₂SO₄, filtered, and evaporated under reduced pressure. The resulting yellow oil was purified by flash column chromatography, eluting with CH₂Cl₂-PE (3:4) → CH₂Cl₂-PE (1:1). The title compound was obtained as a slight yellowish oil that slowly crystallized (3 d) at -18 ˚C, but rapidly crystallized after addition of a seed crystal (3.72 g, 15.6 mmol, 72%). Yellowish powder, mp 35 ˚C. IR (neat): 3119, 2962, 2559, 2369, 2105, 1967, 1766, 1617, 1594, 1523, 1491, 1347, 1198 cm^-¹. ^¹H NMR (300 MHz, CDCl₃): δ = 5.32 (s, 2 H, CH₂N₃), 7.40-7.46 (m, 2 H, ArH), 8.28-8.34 (m, 2 H, ArH). ^¹³C NMR (75 MHz, CDCl₃): δ = 78.96 (CH₂), 121.74 (CH), 125.45 (CH), 145.72, 152.29, 155.04. MS (EI, 70 eV): m/z (%) = 238 (2) [M]⁺, 196 (12), 166 (36), 139 (40), 122 (42), 56 (100), 28 (71). HRMS (EI): m/z calcd for C₈H₆N₄O₅: 238.0338 [M]⁺; found: 238.0319 [M]⁺. Anal. Calcd for C₈H₆N₄O₅: C, 40.35; H, 2.54; N, 23.53. Found: C, 40.25; H, 2.69; N, 23.26.

General Procedure for Azoc Protection with Azidomethyl 4-Nitrophenyl Carbonate (1) and Analytical Data of Compounds 6b,d,g Azidomethyl 4-nitrophenyl carbonate (1, 300 mg, 1.26 mmol, 1.0 equiv) was dissolved in anhyd DMF (3 mL) and cooled to 0 ˚C. Either of compounds 5a-g (1.26 mmol) was dissolved in anhyd DMF (2 mL) in an separate flask and treated with DIPEA (1.39 mmol, 236 µL, 1.1 equiv). The resulting solution was added dropwise, within 2 min, to the solution of 1. The reaction was monitored via TLC (CH₂Cl₂-PE = 3:2). After complete conversion, the reaction mixture was diluted with EtOAc (200 mL) and washed with Na₂CO₃ solution (8 × 50 mL, 2 M). The combined aqueous solutions were re-extracted with EtOAc (30 mL). The extract was washed once with H₂O (10 mL). The combined organic phases were washed with sat. NaCl solution (2 × 50 mL), dried over Na₂SO₄, filtered, concentrated in vacuo and dried at <10^-³ mbar for at least 3 h. Analytical data of compounds 6a and 6f were found to be identical to those in ref. 2.
Azidomethyl 4-Aminobenzylcarbamate (6b) Orange oil which slowly crystallized at -18 ˚C (2 weeks) to give an orange solid with mp 26-28 ˚C. IR (neat): 3423, 3328, 2154, 2098, 1723, 1697, 1617, 1539, 1512 cm^-¹. ^¹H NMR (300 MHz, CDCl₃): δ = 3.68 (s, 2 H, NH₂), 4.27 (d, ^³ J = 5.9 Hz, 2 H, CH₂Ar), 5.05-5.20 (m, 3 H, CH₂N₃, NH), 6.62-6.68 (m, 2 H, ArH), 7.08 (d, ^³ J = 8.3 Hz, 2 H, ArH). ^¹³C NMR (75 MHz, CDCl₃): δ = 44.89 (ArCH₂), 75.32 (CH₂N₃), 115.25, 127.59, 128.72, 129.04, 146.08, 155.18. ESI-HRMS: m/z calcd for C₉H₁₁N₅O₂Na: 244.0805 [M + Na]⁺; found: 244.0802 [M + Na]⁺.
Azidomethyl 5-Hydroxypentylcarbamate (6d) Colorless oil. IR (neat): 3329, 2937, 2864, 2140, 2100, 1705, 1532 cm^-¹. ^¹H NMR (300 MHz, CDCl₃): δ = 1.36-1.48 (m, 2 H, H-3), 1.51-1.65 (m, 5 H, H-2, H-4, OH), 3.24 (q, ^³ J = 6.7 Hz, 2 H, CH₂NH), 3.65 (t, ^³ J = 6.4 Hz, 2 H, CH₂OH), 5.02 (s, 1 H, NH), 5.13 (s, 2 H, CH₂N₃). ^¹³C NMR (75 MHz, CDCl₃): δ = 22.87 (C-3), 29.51, 32.13, 41.02 (C-1), 62.55 (C-5), 75.22 (CH₂N₃), 155.36 (C=O). ESI-HRMS: m/z calcd for C₇H₁₄N₄O₃Na: 225.0958 [M + Na]⁺; found: 225.0964 [M + Na]⁺.
Methyl 4-[(Azidomethoxy)carbonylamino]- N -methyl-pyrrole-2-carboxylate (6g) Compound 6g was purified via flash column chromatography using a gradient of CH₂Cl₂ → CH₂Cl₂-MeOH (99:1).
Orange brownish powder; mp 136-138 ˚C. IR (neat): 3509, 3310, 2958, 2163, 2141, 2112, 1967, 1717, 1679, 1586, 1565 cm^-¹. ^¹H NMR (500 MHz, DMSO-d ₆): δ = 3.72 (s, 3 H, Me), 3.82 (s, 3 H, Me), 5.27 (s, 2 H, CH₂N₃), 6.69 (d, ⁴ J = 1.9 Hz, 1 H, ArH), 7.14 (d, ⁴ J = 1.9 Hz, 1 H, ArH), 9.80 (s, 1 H, NH). ^¹³C NMR (126 MHz, DMSO-d ₆): δ = 36.09, 50.88, 74.88, 107.56, 118.99, 119.47, 121.92, 152.38, 160.54. ESI-HRMS: m/z calcd for C₉H₁₁N₅O₄Na: 276.0703 [M + Na]⁺; found: 276.0716 [M + Na]⁺.

Ammonium 2-[(Azidomethoxy)carbonylamino]ethyl Hydrogenphosphate (6h) Azidomethyl 4-nitrophenyl carbonate (1, 300 mg, 1.26 mmol, 1.0 equiv) and 2-aminoethyl dihydrogen phosphate (177.7 mg, 1.26 mmol, 1.0 equiv) were added to a flask and suspended in DMF (4 mL). DIPEA (4.16 mmol, 707 µL, 3.3 equiv) and H₂O (3 mL) were added. The resulted suspension was sonicated for 2 min and then stirred at 20 ˚C. After 20 min, a clear solution formed. A complete conversion was detected after 30 min via TLC (CH₂Cl₂-PE = 3:2). The reaction mixture was diluted with H₂O (200 mL) and poured on a DEAE-cellulose column [100 g, 20 cm, HCO₃ ^- form, Express-Ion exchanger D, Whatman (Maidstone, UK)]. The column was washed with H₂O (200 mL) and 100 mM NH₄HCO₃ solution (500 mL). The product-containing fractions were combined and the resulting solution concentrated to 20 mL, followed by lyophilization to dryness. The title compound 6h was obtained as a colorless solid (321 mg, 1.25 mmol, 99%). IR (neat): 3210, 2889, 2103, 1705, 1532, 1455 cm^-¹. ^¹H NMR (300 MHz, D₂O): δ = 3.28 (t, ^³ J = 5.3 Hz, 2 H), 3.75-3.83 (m, 2 H), 5.06 (s, 2 H, CH₂N₃). ^¹³C NMR (75 MHz, D₂O): δ = 41.25 (d,
^³ J _P-C2 = 7.8 Hz, C-2), 63.59 (d, ^² J _P-C1 = 5.2 Hz, C-1), 75.71 (CH₂N₃), 157.35 (C=O). ^³¹P NMR (121.5 MHz, D₂O): δ = 1.10. ESI-HRMS: m/z calcd for C₄H₈N₄O₆P: 239.0187 [M]^-; found: 239.0175 [M]^-.

Ammonium 3-[(Azidomethoxy)carbonylamino]-propanoate (6i)
A sample of β-alanine (112.2 mg, 1.26 mmol) was treated as described in ref. 13 but in the presence of a smaller amount of DIPEA (471 µL, 2.77 mmol, 2.2 equiv). The column purification used a higher concentration of NH₄HCO₃ (250 mM) for elution from the DEAE-cellulose. The title compound was obtained as colorless oil (236.2 mg, 1.15 mmol, 91%). IR (neat): 3231, 2965, 2103, 1967, 1701, 1524, 1447, 1409, 1219 cm^-¹. ^¹H NMR (300 MHz, D₂O): δ = 2.44 (t, ^³ J = 6.6 Hz, 2 H), 3.31 (t, ^³ J = 6.6 Hz, 2 H), 5.06 (s, 2 H, CH₂N₃). ^¹³C NMR (75 MHz, D₂O): δ = 34.95, 36.79, 75.62 (CH₂N₃), 157.14, 177.60. ESI-HRMS: m/z calcd for C₅H₇N₄O₄: 187.0473 [M]^-; found: 187.0468 [M]^-.

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