Synthesis of photolabile protecting group (PPG) protected uronic acid building blocks: applications in carbohydrate synthesis with the assistance of a continuous flow photoreactor

Varsha Tiwaria, Adesh Kumar Singha, Priyanka Chaudharya, Peter H. Seebergerb and Jeyakumar Kandasamy*a
aDepartment of chemistry, Indian Institute of Technology (BHU), Varanasi, Uttar Pradesh-221005, India. E-mail:
bMax-Planck-Institute of Colloids and Interfaces, Department of Biomolecular Systems, Am Mühlenberg 1, 14476 Potsdam, Germany

Received 13th August 2019 , Accepted 13th October 2019

First published on 14th October 2019

Photolabile protecting group (PPG) protected uronic acid building blocks were prepared and used for carbohydrate synthesis. Deprotection of the photolabile protecting group was achieved very efficiently by employing a continuous flow photoreactor under neutral conditions. Many conventional protecting groups were found to be stable during the photo-cleavage of the 2-nitrobenzyl group at 355 nm in methanol.


Uronic acid containing polysaccharides including glycosaminoglycans (GAGs), capsular polysaccharides of various bacteria, and marine polysaccharides (Fig. 1) are involved in many biological functions.1,2 The synthesis of structurally well-defined carbohydrates is crucial for understanding the roles of glycans in biological systems.3
image file: c9qo01010f-f1.tif
Fig. 1 Examples of uronic acid containing polysaccharides.

The synthesis of uronic acid containing oligosaccharides is often challenging since the protection and deprotection of carboxylic acids requires special attention. Carboxylic acids are typically protected as methyl esters using hazardous diazomethane and cleaved at the end of oligosaccharide assembly by saponification under basic conditions (pH > 10). These strong basic conditions can sometimes lead to β-elimination or epimerization due to the acidic proton on C5 (Scheme 1).4 On the other hand, protection of carboxylic acids as benzyl esters was also demonstrated in oligosaccharide synthesis5 while their deprotection was effected using metal hydroxides (i.e. saponification)5a,b or Pd/C, H2 (i.e. hydrogenolysis).5c,d

image file: c9qo01010f-s1.tif
Scheme 1 β-Elimination under basic conditions.

Photolabile protecting groups (PPGs) are appealing for organic synthesis because photo-cleavage typically takes place under neutral conditions without any chemical reagents.6 The protection of carboxylic acids using PPG esters may overcome the elimination or epimerization issues during oligosaccharide synthesis (Scheme 2). PPG protected uronic acids have not been explored previously in carbohydrate synthesis.

image file: c9qo01010f-s2.tif
Scheme 2 Cleavage of PPG protected uronic acids.

Photolabile protecting groups are well explored for natural product total syntheses, but their application in oligosaccharide synthesis remains less explored. Protecting group manipulations involving PPGs in batch photochemical carbohydrate syntheses might be low yielding.7 However, continuous-flow photo-reactors overcome the major challenges associated with the use of batch reactors.8 A higher surface to volume ratio and the proximity of the molecule to the UV lamp ensure effective irradiation of large volumes while minimizing transmission versus distance constraints to provide good yields of the desired products. Photolabile protecting groups may be useful for carbohydrate synthesis when continuous flow photo-reactors are employed. The 2-nitro benzyl group is the most widely utilized among the photolabile protecting groups.5 Our primary objective was the synthesis of 2-nitrobenzyl protected uronic acid building blocks and the cleavage of the PPG with the assistance of a continuous flow photoreactor.

As part of our carbohydrate synthesis program,9 we developed an efficient method for the preparation of various uronic acid building blocks using 1-chloro-1,2-benziodoxol-3(1H)-one and TEMPO at room temperature under neutral conditions.8a Using this method, we prepared various functionalized uronic acids and reacted them with 2-nitrobenzyl bromide in the presence of potassium bicarbonate and tetra-butylammonium iodide (Scheme 3).

image file: c9qo01010f-s3.tif
Scheme 3 Synthesis of PPG protected uronic acids.

Results and discussion

Continuous flow photoreactor construction

The continuous flow photo-reactor (CFPR) was constructed with the help of M/s Lelesil Innovative Systems, Mumbai, India. The reactor (Fig. 2) is composed of a main photo-reactor unit within a photo-safety cabinet, a digital lamp controller unit, a chiller and a peristaltic pump. The main unit of the system is composed of a stand, a quartz jacket with an inlet and an outlet, a flexi coil (height 10 mm × 100 mm, diameter 4 mm) which is made up of a transparent FEP (Fluorinated Ethylene Propylene) tube obtained from M/s BOLA, Germany and a 250 W medium pressure mercury vapour lamp (MPMVL). This main unit is placed in a safety cabinet which is equipped with an exhaust fan and a LED window. The inlet and outlet of the quartz jacket are connected to the chiller for cooling. The chiller circulates cold water (∼5–8 °C) into the quartz jacket in order to neutralize the temperature generated by the lamp. The peristaltic pump inlet is connected to reagent bottle B (to store the solution to be photolyzed) and the outlet is connected to the flexi FEP coil. The upper side of the FEP tube is connected to another reagent bottle A to receive the photolyzed solution. The medium pressure mercury lamp is placed in the quartz jacket and connected to the digital lamp controller unit.
image file: c9qo01010f-f2.tif
Fig. 2 Schematic diagram of the continuous flow photoreactor.

Cleavage of PPG in uronic acids using the continuous flow photoreactor

To find the best conditions, 2-nitrobenzyl protected α-methyl tri-O-benzyl glucuronic acid 1a was subjected to photolysis using the continuous flow photo-reactor (Table 1). The reaction was performed in polar solvents including methanol, THF, DCM, DMF, acetonitrile and 1,4-dioxane (Table 1, entries 1–6). A 0.005 M solution of the substrate was prepared in an appropriate solvent (50 mL) and added to reactor container B and circulated with the help of the peristaltic pump to the flexi coil. The peristaltic pump was maintained at 50 RPM which requires approximately 3 minutes to complete one cycle of irradiation. The reaction yield was analyzed under irradiation for 3, 6 and 9 cycles respectively in methanol (Table 1, entries 7–9). The reaction was found to be most efficient in six cycles (=18 min) which yielded 92% of the desired product resulting in the complete cleavage of PPG (Table 1, entry 8). In a batch reactor setup, only 29% desired product was obtained after one hour while 64% product was obtained after two hours (Table 1, entries 10 and 11). In fact, there was only slight improvement in the yield in the batch reactor even after 4 h (Table 1, entry 12). Nevertheless, it is worthwhile to mention that the PPG protected uronic acid 1a was found to be very stable in indoor lighting and in sunlight.10
Table 1 Optimization of photo-deprotection using flow and batch reactorsa,b

image file: c9qo01010f-u1.tif

Entry Solvent No. of cycles Time (min) Yield (%)
a 0.005 molar solution of the PPG protected uronic acid was prepared in different solvents and stored in reactor B.b Isolated yield.
1 CH3OH 1 3 60
2 THF 1 3 20
3 CH2Cl2 1 3 >5
4 DMF 1 3 19
5 CH3CN 1 3 34
6 1,4-Dioxane 1 3 22
7 CH3OH 3 9 78
8 CH3OH 6 18 92
9 CH3OH 9 27 89
10 CH3OH Batch reactor 60 29
11 CH3OH Batch reactor 120 64
12 CH3OH Batch reactor 240 70

With the optimized conditions in hand, different PPG protected glucuronic acid, mannuronic acid, and galacturonic acid equipped with different conventional protecting groups were subjected to photocleavage using the continuous flow photo-reactor (Table 2). The PPG was selectively cleaved in all the substrates as the desired products were obtained in quantitative yields. Other protecting groups such as benzyl, benzoyl, acetyl, acetonide, 4-bromobenzyl,11 2-naphthylmethyl and carbamate were found to be very stable during the photo-cleavage of the 2-nitrobenzyl group.

Table 2 Photo-deprotection of various uronic acid building blocksa,b
a 0.005 molar solution of the PPG protected uronic acid was subjected to photolysis with a flow rate of 50 RPM.b Isolated yield.
image file: c9qo01010f-u2.tif

The deprotection protocol was applied to a disaccharide containing two photolabile protecting groups (Scheme 4). Still, the deprotection proceeds for the disaccharide as efficiently as for monosaccharides.

image file: c9qo01010f-s4.tif
Scheme 4 Cleavage of photolabile protecting groups in disaccharides.

To investigate the compatibility of PPGs with glycosylation conditions, the PPG protected glycosyl imidate 1aa was used to glycosylate sugar and non-sugar acceptors (3a–c) in the presence of tris(pentafluorophenyl)borane (Table 3).8c These reactions yielded the desired glycosides (4a–c) in good yields as the PPG remained intact under these glycosylation conditions. The PPG in the glycosides 4a–c were cleaved using the flow reactor to provide uronic acids 5a–c in 90–94% yields. Furthermore, we investigated the glycosylation of benzyl alcohol with a PPG protected thioglycoside donor (1h) in the presence of NIS/TfOH in DCM (Scheme 5). To our delight, the desired glycosylated product 4d was obtained in 52% yield. The compound 4d was subsequently subjected to photo-deprotection in a flow reactor under the optimized conditions to obtain the uronic acid 5d in 84% yield.

image file: c9qo01010f-s5.tif
Scheme 5 Glycosylation with a thioglycoside donor followed by photo-deprotection.
Table 3 Glycosylation followed by photo-deprotection of various uronic acid building blocksa,b

image file: c9qo01010f-u3.tif

R–OH (acceptor) Glycosylation product Photolyzed product
a Reaction conditions: Donor: 1aa (0.5 mmol), acceptor: 3a and 3b (3 equiv.), 3c (1.2 equiv.), CH2Cl2 (8 mL), 10 mol% B(C6F5)3; mol. sieves (4 Å).b Isolated yield.
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image file: c9qo01010f-u10.tif image file: c9qo01010f-u11.tif image file: c9qo01010f-u12.tif


The use of a photolabile protecting group in uronic acid building blocks was investigated. The photolabile protecting group can be selectively cleaved in an excellent yield in the presence of other protecting groups such as acetate, benzoate, acetonide, halobenzyl, 2-naphthyl methyl and carbamate. This protocol should prove to be useful for the synthesis of complex oligosaccharides.

Conflicts of interest

There are no conflicts to declare.


We thank the Max-Planck Society (Indo-Max Planck partner group) and DST-India (DST/INT/MPG/P-09/2016 to J. K.) for generous financial support of the project. J. K. acknowledges the Central Instrumentation Facility Centre (CIFC)-IIT (BHU) for the NMR facilities.


  1. (a) G. O. Aspinall, The polysaccharides, Academic Press, New York, 1985 Search PubMed; (b) J. D. C. Codée, A. E. Christina, M. T. C. Walvoort, H. S. Overkleeft and G. A. van der Marel, Top. Curr. Chem., 2011, 301, 253–289 CrossRef PubMed; (c) L. J. Van den Bos, J. D. C. Codée, R. E. J. N. Litjens, J. Dinkelaar, H. S. Overkleeft and G. A. van der Marel, Eur. J. Org. Chem., 2007, 3963–3976 CrossRef CAS.
  2. (a) M. Mende, C. Bednarek, M. Wawryszyn, P. Sauter, M. B. Biskup, U. Schepers and S. Brase, Chem. Rev., 2016, 116, 8193–8255 CrossRef CAS PubMed; (b) M. Petitou, J. C. Lormeau and J. Choay, Nature, 1991, 350, 30–33 CAS; (c) S.-K. Kim, Marine Glycobiology: Principles and Applications, CRC Press, Taylor & Francis Group, New York, 2017 Search PubMed; (d) D. M. Weinberger, K. Trzcinski, Y. J. Lu, D. Bogaert, A. Brandes, J. Galagan, P. W. Anderson, R. Malley and M. Lipsitch, PLoS Pathog., 2009, 5, e1000476 CrossRef PubMed.
  3. (a) A. Varki, Essentials of Glycobiology, Cold Spring Harbor Laboratory Press, New York, USA, 1999 Search PubMed; (b) T. K. Lindhorst, Essentials of Carbohydrate Chemistry and Biochemistry, Wiley-Vch Verlag GmbH & Co. Weinheim, 2008 Search PubMed; (c) P. H. Seeberger and D. B. Werz, Nature, 2007, 446, 1046–1051 CrossRef CAS PubMed; (d) P. H. Seeberger and D. B. Werz, Nat. Rev. Drug Discovery, 2005, 4, 751–763 CrossRef CAS PubMed; (e) M. C. Galan, D. Benito-Alifonso and G. M. Watt, Org. Biomol. Chem., 2011, 9, 3598–3610 RSC.
  4. (a) P. Sjolin and J. Kihlberg, J. Org. Chem., 2001, 66, 2957–2965 CrossRef CAS PubMed; (b) G. Tiruchinapally, Z. J. Yin, M. El-Dakdouki, Z. Wang and X. F. Huang, Chem. – Eur. J., 2011, 17, 10106–10112 CrossRef CAS PubMed; (c) I. N. Bemiller and G. V. Kumari, Carbohydr. Res., 1972, 25, 419–428 CrossRef; (d) M. J. H. Keijbets, Carbohydr. Res., 1974, 33, 359–362 CrossRef CAS.
  5. (a) X. Lu, M. N. Kamat, L. Huang and X. Huang, J. Org. Chem., 2009, 74, 7608–7617 CrossRef CAS PubMed; (b) X. Dai, W. Liu, Q. Zhou, C. Cheng, C. Yang, S. Wang, M. Zhang, P. Tang, H. Song, D. Zhang and Y. Qin, J. Org. Chem., 2016, 81, 162–184 CrossRef CAS PubMed; (c) E. R. Bowkett, J. R. Harding, J. L. Maggs, B. K. Park, J. A. Perrie and A. V. Stachulski, Tetrahedron, 2007, 63, 7596–7605 CrossRef CAS; (d) P. Chassagne, C. Fontana, C. Guerreiro, C. Gauthier, A. Phalipon, G. Widmalm and L. A. Mulard, Eur. J. Org. Chem., 2013, 4085–4106 CrossRef CAS.
  6. (a) P. Klán, T. Šolomek, C. G. Bochet, A. Blanc, R. Givens, M. Rubina, V. Popik, A. Kostikov and J. Wirz, Chem. Rev., 2013, 113, 119–191 CrossRef PubMed; (b) H. T. Yu, J. B. Li, D. D. Wu, Z. J. Qiu and Y. Zhang, Chem. Soc. Rev., 2010, 39, 464–473 RSC; (c) C. G. Bochet, J. Chem. Soc., Perkin Trans. 1, 2002, 125–142 CAS; (d) P. F. Wang, Asian J. Org. Chem., 2013, 2, 452–464 CrossRef CAS.
  7. (a) J. P. Knowles, L. D. Elliott and K. I. Booker-Milburn, Beilstein J. Org. Chem., 2012, 8, 2025–2052 CrossRef CAS PubMed.
  8. (a) Y. H. Su, N. J. W. Straathof, V. Hessel and T. Noel, Chem. – Eur. J., 2014, 20, 10562–10589 CrossRef CAS PubMed; (b) B. D. A. Hook, W. Dohle, P. R. Hirst, M. Pickworth, M. B. Berry and K. I. Booker-Milburn, J. Org. Chem., 2005, 70, 7558–7564 CrossRef CAS PubMed; (c) K. Gilmore and P. H. Seeberger, Chem. Rec., 2014, 14, 410–418 CrossRef CAS PubMed.
  9. (a) V. Tiwari, V. N. Badavath, A. K. Singh and J. Kandasamy, Tetrahedron Lett., 2018, 59, 2511–2514 CrossRef CAS; (b) A. K. Singh, V. Tiwari, K. B. Mishra, S. Gupta and J. Kandasamy, Beilstein J. Org. Chem., 2017, 13, 1139 CrossRef CAS PubMed; (c) K. B. Mishra, A. K. Singh and J. Kandasamy, J. Org. Chem., 2018, 83, 4204 CrossRef CAS PubMed; (d) A. K. Singh and J. Kandasamy, Org. Biomol. Chem., 2018, 16, 5107–5112 RSC; (e) K. B. Mishra and J. Kandasamy, Asian J. Org. Chem., 2019, 8, 549–554 CrossRef.
  10. We observed only less than 5% decomposition of PPG protected uronic acid 1a after 48 hours under indoor lighting (5 W white LED was used). To check the effect of sunlight, an experiment was performed in open sunlight during the month of September (under North-East Indian climatic conditions, room temperature (33–35 °C)). The PPG protected uronic acid 1a (100 mg) was dissolved in methanol (2 mL) and was kept under sunlight for a period of 1 hour. Only less than 5% decomposition of 1a was observed.
  11. O. J. Plante, S. L. Buchwald and P. H. Seeberger, J. Am. Chem. Soc., 2000, 122, 7148–7149 CrossRef CAS.


Electronic supplementary information (ESI) available. See DOI: 10.1039/c9qo01010f
These authors contributed equally to this work.

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