Fmoc quantitation

The linker was synthesized from Merrifield resin or carboxypolystyrene and a DMT-protected diol (Scheme 2). The cleavage of the resulting DMT-protected Unker allowed for UV quantitation of the resin loading [22]. The linker can also be installed directly from the diol [23]. The loading can be determined by Fmoc quantitation [24]. 

This is a very important and well tested method for the quantitative determination of loading of Fmoc protected compounds particularly that of Fmoc (fluorenylmethoxycarbonyl) amino acids on solid support. Fmoc groups can... 

N -Fmoc serine benzyl ester 2, which could be prepared as shown or purchased commercially, was smoothly converted to the crystalHne O-methylthiomethyl (MTM) ether 3 in high yield via a Pummerer-Hke reaction using benzoyl peroxide and dimethyl sulfide in acetonitrile [39]. This common intermediate was used to synthesize both 5 and 8 [40]. Both Ogilvie [41] and Tsantrizos [42] had reported that I2 was an effective activator with similar MTM ether substrates. The H promoted nucleosidation reaction between O-MTM ether 3 and bis-silylated thymine 4 produced the nucleoamino acid 5 in 60% isolated yield (100% based on recovered 3). Hydrogenolytic deprotection of the benzyl ester with H2, Pd/C in MeOH gave the thymine-containing nucleoamino acid 6 in quantitative yield. 

OS 16] ]R 5] ]P 13] Using continuous flow in an electroosmotic-driven micro reactor gave a quantitative yield of the dipeptide in only 20 min (600 V for Dmab-/ -alanine 700 V for the Fmoc ester) [5, 88]. Batch synthesis under the same conditions gave only a 40-50% yield [5] (46% in [5]), needing 24 h. 

OS 18] ]R 5] ]P 13] (adapted) The same was done using Fmoc-L-y9-homo-p-chlorophenylalanine [5]. Batch, 36% micro reactor, quantitative in 20 min. 

Acidolytic treatment using DCM hexafluoroisopropanoI (1 1) for 2 h at ambient temperature afforded the hydroxamic acid in only 45% yield. However, it is worth noting that the tethered Fmoc-N(Pr)-<9-2-chIorotrityI polystyrene, on treatment with similar acidolytic cocktail effected quantitative release of Fmoc-N(Pr)-OH. 

An alternative approach is the cleavage of a UV-active protecting group from the resin, such as the widely used Fmoc Test. The quantitation of the 9-fluorenyl-methyloxycarbonyl (Fmoc) protecting group for amines is used in SPPS as an indirect method to determine the extent of a peptide coupling reaction. Similar approaches have also been recently reported for the quantitation of supported thiols [151, 154] and have also been the subject of an excellent review [148]. 

TH-0-acetyl-2-azido-2-deoxy-a-D-galactopyranosyl)-L-threonine tert-Butyl Ester 22. A solution of Fmoc-Thr(aAc3GalN3)-OtBu 20 (215 mg, 0.3 mmol) in freshly distilled morpholine (2 mL) is stirred at room temperature for 20 min. After concentration in vacuo and codistillation with toluene (twice 3 mL) in vacuo, 22 is obtained quantitatively. It is immediately used in the following reaction. 

After the reaction times given in Scheme 38, the respective couplings were quantitative according to chloranil and the Kaiser ninhydrin test and/or HPLC of small cleavage. The peak area of the crude Fmoc-deprotected dodecapeptide was 90% (HPLC k=214nm). The pure cyclic peptide was obtained in an overall yield of 31%. Cyclosporin O was synthesized by a similar procedure.129 ... 

A report involving the solid-phase preparation of Fmoc-protected t t[CH2—S] pseudodipeptides revealed that the expected stereochemical patterns did not hold.147 It is known that the conversion of an a-bromo acid into the a-thioacid does not always occur with quantitative inversion of chirality, since it is side-chain dependent.146] With side chains such as benzyl (Phe) or butyl (Leu, lie), exceptions are not expected. But when the Fmoc-protected amino thiol 26 was condensed to a support-bound bromo acid 27, as shown in Scheme 8, the (S,R)-product 28 was obtained instead of the expected (S,S)-isomer. 

The combination of very mild coupling and deprotection protocols provides the flexibility needed in order to incorporate a variety of side-chain-protecting groups, cleavable by acid (Boc), base (Fmoc), Pd(II) (Aloc)J43 or nucleophiles (Dde)J441 Linear precursors of such templates with four orthogonally protected lysine residues are generally obtained in yields around 80% and cyclization proceeds quickly at high dilution almost quantitatively. 

Synthesis of octreotide and derivative thereof can be carried out by two methods. The first method is synthesized initially by fragment condensation solution phase procedures. The synthetic process of octreotide has been described by Bauer et al. (1). The second method is the synthese by solid-phase procedures. Edward et al. (2) isolated side chain protected octreotide with a total yield of 14% by cleaving the protected peptide from the resin with threoninol. Arano et al. (3) carried out another solid phase method for octreotide, and produced it in overall 31.8% yield based on the starting Fmoc-Thr(tBu)-ol-resin. The basic difference from the other procedures already described is that the introduction of the threoninol is carried out upon the protected peptidic structure (resin-free), which, when appropriately activated, leads quantitatively and without needing to make temporary protections upon the threoninol, to the protected precursor of octreotide, which in turn, with a simple acid treatment leads to octreotide with very high yields. 

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