The benzyl esters 1 ± 3 of oligo[(R)-3-hydroxybutanoic acids] (OHB) containing 2, 16, or 32 HB units were coupled at the hydroxy terminus with arginine (by esterification with carbodiimide), with glucose (by acetalization with glucosyl trichloroacetimidate), and with 7-(dimethylamino)coumarin-4-acetic acid and biotin (by amide formation through a glycine linker) to give, after deprotection(s), the corresponding labelled OHB acids 7 ± 9, 12, 13, 25, 26, 33, and 34 (Schemes 1, 4,and 5). The respective novel 16-and 32mer derivatives exhibit distinct water solubility (Table ) or may be detected (in minute amounts) by fluorescence spectroscopy, properties required for biochemical investigations.
Helvetica Chimica Acta ± Vol. 81 (1998) CH 2 Cl 2 , to give the conjugates 4 (62%), 5 (64%), and 6 (36%). By using the N-Fmoc-N w -Pmc-protected arginine under otherwise similar conditions, the formation of several by-products was observed. Coupling with the N-Z-N w -NO 2 -protected arginine derivative yielded the desired product in good yield (not shown), however, the subsequent deprotection did not work in our case. Hydrogenolytic debenzylation of compounds 4 ± 6 with H 2 and Pd/C produced the fully deprotected compounds 7 ± 9 in excellent yields.
Coupling of an OHB with a carbohydrate moiety was achieved by following the glycosylation procedure introduced by Schmidt [16]. Reaction of the hydroxy esters 1 and 2 with tetra-O-benzyl-d-glucosyl trichloroacetimidate, readily available in one step from tetra-O-benzyl-d-glucose, gave the fully benzyl-protected conjugates 10 (93%) and 11 (82%), respectively (Scheme 1). We were not able to separate the anomers by flash chromatography, but for the intended studies, the configuration has no significance. Hydrogenolytic cleavage of the benzyl ester and ethers in 10 and 11 with H 2 and Pd/C gave the desired products 12 (88%) and 13 (77%), respectively.
The water solubility of the OHB conjugates 8, 9, and 13 was compared with those of their precursors 2 and 3 (Table ). Thus, the solubility of the 16mer was enhanced from Scheme 1. Synthesis of Water-Solubilized HB Derivatives a) Tri-Z-l-arginine, DCC, DMAP, CH 2 Cl 2 , 08. b) H 2 , Pd/C, CF 3 CH 2 OH, r.t. c) 2,3,4,6-Tetra-O-benzyl-dglucosyl trichloroacetimidate, BF 3 ´Et 2 O, CH 2 Cl 2 , 08. DCC dicyclohexylcarbodiimide, DMAP 4-(dimethylamino)pyridine Helvetica Chimica Acta ± Vol. 81 (1998) 2417 7 ) Attempts to prepare the acid chloride of 7-(dimethylamino)coumarin-4-acetic acid with oxalyl or thionyl chloride failed. 8 ) The use of various bases, solvents, and reaction temperatures did not lead to better results. 9 ) Hydrogenolysis with H 2 and Pd/C gave OHBs with free carboxy and hydroxy function, resulting from benzyl-ester and SÀO bond cleavage. Catalytic transfer hydrogenolysis (Pd/C, cyclohexa-1,4-diene, DMF) gave OHBs with a,b-unsaturated carbonyl functionalities, resulting from a b-elimination of dansyl sulfate. 10 ) Because of the higher nucleophilicity of the amino group and the enhanced stability of the newly formed amide bond, the coupling reaction was expected to work better, what it actually did. a ) Based on material isolated from SiO 2 chromatography. b ) BOP-Cl bis(2-oxooxazolidin-3-yl)phosphinic chloride. Scheme 3. Synthesis of Fluorescent OHB Derivatives a) 7-(Dimethylamino)coumarin-4-acetic acid, 2-chloro-1-methylpyridinium iodide, Et 3 N, CH 2 Cl 2 , MeCN, r.t. b) Cyclohexa-1,4-diene, Pd/C, DMF, r.t. c) 5-(Dimethylamino)naphthalene-1-sulfonyl chloride, DMAP, 508, ClCH 2 CH 2 Cl. Helvetica Chimica Acta ± Vol. 81 (1998) 2419 Scheme 4. Synthesis of HB Derivatives Linked to a Coumarin via a Glycine Spacer a) DCC, DMAP, CH 2 Cl 2 , 08. b) 1. CF 3 COOH, CH 2 Cl 2 , 08, 2. Et 3 N, CHCl 3 , 08; 3. 7-(dimethylamino)coumarin-4-acetic acid, EDC, HOBt, DMF, CHCl 3 , 08 3 r.t. c) Cyclohexa-1,4-diene, Pd/C, DMF, r.t. or H 2 , Pd/BaSO 4 , CF 3 CH 2 OH, r.t. EDC N-[3-(dimethylamino)propyl]-N'-ethylcarbodiimide hydrochloride 11 ) For comparison: the 32mer HB with free carboxy and hydroxy function has a UV absorption maximum at 214 nm and a molar extinction coefficient e of 3326 m À1 cm À1 (AE 2.5%) in CF 3 CH 2 OH. The 376-nm coumarin absorption does not give rise to a Cotton effect of compound 26.