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Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010Tert-BuNH 2 as an Efficient Reagent for the Deprotection ofFmoc Protected Amino AcidsArmin Arabanian 1,2 and Saeed Balalaie 11 Peptide Chemistry Research Center, K.N. Toosi University of Technology, Tehran,15875-4416, Iran; 2 Tofigh Daru Res. & Eng. Co., 61 st St. km 18 Karaj Highway,Tehran, 37515-375, IranIntroductionThe protection and deprotection of the amine group is essential for the condensation ofα-amino acids via amide bond in a defined sequential order. All amino protecting groupsare fundamentally suitable for masking the α-amino group of amino acids, but it is veryimportant that the repetitive steps proceed rapidly in high yields and with minimal sidereactions to prevent formation of by-products. N α Deprotection is one of the major steps inSPPS with consideration of orthogonality. The Fmoc/tBu method was introduced byAtherton and Meienhofer and is based on an orthogonal protecting group strategy, using thebase labile Fmoc group for protection of the α-amino group and acid labile side-chainprotecting groups and resin-linkage agents [1]. In Fmoc/tBu SPPS, a 20% V/V solution ofpiperidine in DMF is used as an efficient condition for Fmoc-deprotection. The detailsabout the mechanism were clarified. It was shown that, in some cases the standardtreatment with 20% piperidine in DMF may not always be effective and need a strongerbase such as DBU. Piperidine is not only an expensive reagent, but also a controlledsubstance according to the 92/109/EC recommendation, and also there are some reportsabout the effect of piperidine on mutation and DNA destruction. According to thesedrawbacks, Leondiadis reported using of 5% piperidine for Fmoc-deprotection [2].Results and DiscussionFinding a suitable reagent for Fmoc-deprotection is an interesting subject in peptidesynthesis chemistry. In this work, we report experimentally a simple procedure for theN α deprotection using t-BuNH 2 to remove Fmoc in SPPS. Recently, t-BuNH 2 was used forthe selective cleavage of carbamates [3]. To examine the possibility of using t-BuNH 2solution, different percentages of it were used. We applied 25, 10 and 5% t-BuNH 2 solutionfor the Fmoc-deprotection from different Fmoc-protected amino acids. Meanwhile, wechecked the rate of Fmoc-deprotection and the sample was collected after 2, 5, 10 and 20min. The protected amino acids on 2-chlorotrityl chloride resin were selected as startingmaterials and removal of Fmoc group was controlled by using UV absorption at 301 nm.The details about Fmoc-deprotection from two amino acids (Leu and Met) which weresupported on resin were summarized in Tables 1 and 2.According to these data, t-BuNH 2 in DMF provided the same result like piperidine forthe Fmoc-deprotection. For example, we tested the use of 5% t-BuNH 2 solution and after20 min, more than 90% of the Fmoc group was removed. The Fmoc-deprotection using t-BuNH 2 is slower than piperidine, but when 25% t-BuNH 2 was used, the same results asTable 1. Fmoc-deprotection using t-BuNH 2 for Fmoc-Leu-ResinBasePiperidine/DMFt-Butylamine/DMFFmoc-deprotection (%)T=2 min T=5 min T=10 min T=20 min5% 53.8 83.6 92.8 97.510% 77.6 86.2 89.7 98.225% 81.6 95.1 98.6 100.05% 30.0 51.4 65.2 92.810% 31.3 64.4 76.0 95.225% 60.8 85.1 90.4 100.050