A rapid coupling protocol for the synthesis of peptide nucleic acids

Summary With the current interest in anti-sense and anti-gene technologies, an efficient, fast and less toxic synthesis protocol would be advantageous for the oligomerisation of Peptide Nucleic Acids (PNA). Most of the methods currently in use for thet-Boc synthesis of PNA's use TFA/m-cresol, pyridine, piperidine and capping reagents. In this work, a rapid synthesis protocol has been adapted from an earlier published peptide synthesis method allowing a reduction in cycle time from around 30 min down to 16 min. By utilising quantitative deprotection with 100% TFA, a coupling time of 10 min and a four-fold excess of monomer, this synthesis protocol has been used to synthesise a number of PNA's incorporating all four nucleotides of varying sequence, up to 17 residues in length.     

Acid-promoted synthesis of per-O-sulfated fucooligosaccharides related to fucoidan fragments

The synthesis of per-O-sulfated derivatives of di-, tetra-, hexa-, octa-, dodeca-, and hexadecafucosides related to natural fucoidans of different types has been performed with the use of previously reported acid-promoted protocol for per-O-sulfation of polyols by SO₃ complexes.² During the treatment of (1→3)-linked oligofucosides under these conditions with the promotion by TfOH, the unusual rearrangement of the reducing pyranose residue into furanose one was observed. To avoid the formation of rearrangement by-products, the use of a series of strong acids as promoters of sulfation of large oligofucosides was studied and the improved protocol was developed based on the use of TFA instead of TfOH. The efficiency of the new method was demonstrated by the syntheses of per-O-sulfated derivatives of dodeca- and hexadecafucosides. The described method of O-sulfation opens access to the preparation of the oligosaccharides related to fucoidan fragments and their per-O-sulfated derivatives interesting for elucidation of the relationship between their structure and biological activity.     

Side reactions in the SPPS of Cys-containing peptides

Alkylation of sensitive amino acids during synthesis of biologically important peptides is a common and well-documented problem in Fmoc-based strategy. Herein, we probed for the first time an unexpected S-alkylation of Cys-containing peptides that occur during the final TFA cleavage of peptides from the Wang solid support. Through a battery of approaches (NMR, UV and LC–MS) the formed by-product was assigned as the alkylation of the cysteine sulfydryl group by the p-hydroxyl benzyl group derived from the acidic Wang linker decomposition. Factors affecting this side reaction were monitored and a protocol that minimizes the presence of the by-product is reported.     

Microwave assisted,one-pot synthesis of 5-nitro- 2-aryl substituted-1H-benzimidazole libraries: Screening in vitro for antimicrobial activity

The efficient and rapid synthesis of 5-nitro-2-aryl substituted-1H-benzimidazole libraries (1a-1j) has been established. Thus, both microwave and conventional cyclo-condensation of 4-nitro ortho-phenylenediamine with various phenoxyacetic acids were carried out in the presence of HCl catalyst. The microwave synthesis route afforded advantages, such as good to excellent yields, shorter reaction time (2.5–3.5?min), readily available starting material, and simple purification procedure, which distinguish the present protocol from other existing methods used for the synthesis of benzimidazole libraries. Bioassay indicated that all the compounds showed in vitro antimicrobial activity against Vancomycin resistant enteroccoccus, Staphylococcus aureus, Micrococcus, Bacillus subtilis (gram-positive bacteria) and Shigella dysentery, Escherichia coli (gram-negative bacteria) and Candida albicans, Aspergillus niger, Penicillium (fungi). The minimum inhibitory concentration (MIC) was determined for test compounds as well as for reference standards.     

Environmental Risk Assessment of Trifluoroacetic Acid

The Montreal Protocol was developed in 1987 in response to concerns that the chlorofluorocarbons (CFCs) were releasing chlorine into the stratosphere and that this chlorine was causing a depletion of stratospheric ozone over Antarctica. This international agreement called for a phase out of these CFCs. Industry initiated a major effort to find replacements that are safe when properly used and safe to the environment. The toxicology and environmental fate of these first generation replacements has been studied extensively. It was determined that the new substances break down in the environment to give predominantly carbon dioxide, water and inorganic salts of chlorine and fluorine. The only exception is that some substances also break down to yield trifluoroacetic acid (HTFA), a substance resistant to further degradation. Recognizing this, industry embarked on a research and assessment program to study the potential effects of trifluoroacetate (TFA) on the environment and to investigate possible degradation pathways. The results of these recently completed studies are summarized below and described in further detail in this paper. Trifluoroacetic acid is a strong organic acid with a pKa of 0.23. It is miscible with water and its low octanol/water partition coefficient (log P_ow=?2.1) indicates no potential to bioaccumulate. Industrial use is limited and environmental releases are very low. Some additional TFA will be formed from the breakdown of a few halogenated hydrocarbons, most notably HFC-134a (CF₃CH₂F), HCFC-124 (CF₃CHFCl), and HCFC-123 (CF₃CHCl₂). As these substances have only been produced in limited commercial quantities, their contribution to environmental levels has been minimal. Surprisingly, environmental measurements in many of diverse locations show existing levels of 100 to 300?ng·l^?1 in water with one site (Dead Sea) having a level of 6400?ng·l^?1. These levels cannot be accounted for based on current atmospheric sources and imply a long-term, possibly pre-industrial source. Generally, soil retention of TFA is poor although soils with high levels of organic matter have been shown to have a greater affinity for TFA when contrasted to soils with low levels of organic matter. This appears to be an adsorption phenomenon, not irreversible binding. Therefore, TFA will not be retained in soil, but will ultimately enter the aqueous compartment. Modeling of emission rates and subsequent conversion rates for precursors has led to estimates of maximum levels of TFA in rain water in the region of 0.1?µ·^?1 in the year 2020. TFA is resistant to both oxidative and reductive degradation. While there had been speculation regarding the possibility of TFA being degraded into monofluoroacetic acid (MFA), the rate of breakdown of MFA is so much higher than for TFA that any MFA formed would rapidly degrade. Therefore, there would be no buildup of MFA regardless of the levels of TFA present in the environment. Although highly resistant to microbial degradation, there have been two reports of TFA degradation under anaerobic conditions. In the first study, natural sediments reduced TFA. However, even though this work was done in replicate, the investigators and others were unable to reproduce it in subsequent studies. In the second study, radiolabeled TFA was removed from a mixed anaerobic in vitro microcosm. Limited evidence of decarboxylation has also been reported for two strains of bacteria grown under highly specific conditions. TFA was not biodegraded in a semi-continuous activated sludge test even with prolonged incubation (up to 84 days). TFA does not accumulate significantly in lower aquatic life forms such as bacteria, small invertebrates, oligochaete worms and some aquatic plants including Lemna gibba (duckweed). Some bioaccumulation was observed in terrestrial higher plants, such as sunflower and wheat. This result appeared to be related to uptake with water and then concentration due to transpiration water loss. When transferred to clean hydroponic media, some elimination of TFA was seen. Also, more than 80% of the TFA in leaves was found to be water ex-tractable, suggesting that no significant metabolism of TFA had occurred. At an exposure level of 1200?mg·l^?1 of sodium trifluoroacetate (NaTFA) — corresponding to 1000?mg·l^?1 HTFA — no effects were seen on either Brachy-danio rerio (a fish) or Daphnia magna (a water flea). With duckweed, mild effects were seen on frond increase and weight increase at the same exposure level. At a concentration of 300?mg·l^?1 no effects were observed. Toxicity tests were conducted with 11 species of algae. For ten of these species the EC50 was greater than 100?mg·l^?1. In Selenastrum capricornutum the no-effect level was 0.12?mg·l^?1. At higher levels the effect was reversible. The reason for the unique sensitivity of this strain is unknown, but a recovery of the growth rate was seen when citric acid was added. This could imply a competitive inhibition of the citric acid cycle. The effect of TFA on seed germination and plant growth has been evaluated with a wide variety of plants. Application of NaTFA at 1000?mg·l^?1 to seeds of sunflower, cabbage, lettuce, tomato, mung bean, soy bean, wheat, corn, oats and rice did not affect germination. Foliar application of a solution of 100?mg·l^?1 of NaTFA to field grown plants did not affect growth of sunflower, soya, wheat, maize, oilseed rape, rice and plantain. When plantain, wheat (varieties Katepwa and Hanno) and soya were grown in hydroponic systems containing NaTFA, no effects were seen on plantain at 32?mg·l^?1, on wheat (Katepwa) and soya at 1?mg·l^?1, or on wheat (Hanno) at 10?mg·l^?1; some effects on growth were seen at, respectively, 100?mg·l^?1, 5?mg·l^?1, 5?mg·l^?1, and 10?mg·l^?1 and above. TFA is not metabolized in mammalian systems to any great extent. It is the major final metabolite of halothane, HCFC-123 and HCFC-124. The half-life of TFA in humans is 16 hours. As expected, the acute oral toxicity of the free acid is higher than the one of the sodium salt. The inhalation LC₅₀ (2 hour exposure) for mice was 13.5?mg·l^?1 (2900?ppm) and for rats it was 10?mg·l^?1 (2140?ppm). Thus, TFA is considered to have low inhalation toxicity. The irritation threshold for humans was 54?ppm. As one would expect of a strong acid, it is a severe irritant to the skin and eye. When conjugated with protein, it has been shown to elicit an immunolog-ical reaction; however, it is unlikely that TFA itself would elicit a sensitization response. Repeat administration of aqueous solutions have shown that TFA can cause increased liver weight and induction of peroxisomes. Relative to the doses (0.5% in diet or 150?mg·kg^?1·day^?1 gavage) the effects are mild. In a series of Ames assays, TFA was reported to be non-mutagenic. Its carcinogenic potential has not been evaluated. Although TFA was shown to accumulate in amniotic fluid following exposure of pregnant animals to high levels of halothane (1200?ppm), no fetal effects were seen. Likewise, a reproduction study that involved exposure of animals to halothane at levels up to 4000?ppm for 4 hours per day, 7 days per week, resulted in no adverse effects. Given the high levels of halothane exposure, it is unlikely that environmental TFA is a reproductive or developmental hazard. Overall the toxicity of TFA has been evaluated in stream mesocosms, algae, higher plants, fish, animals and humans. It has been found to be of very low toxicity in all of these systems. The lowest threshold for any effects was the reversible effect on growth of one strain of algae, Selenastrum capricornutum, which was seen at 0.12?mg·l^?1. There is a 1000-fold difference between the no-effect concentration and the projected environmental levels of TFA from HFCs and HCFCs (0.0001?mg·l^?1). Based on available data, one can conclude that environmental levels of TFA resulting from the breakdown of alternative fluorocarbons do not pose a threat to the environment.     

In Situ Neutralization in Boc-chemistry Solid Phase Peptide Synthesis

Simple, effective protocols have been developed for manual and machine-assisted Boc-chemistry solid phase peptide synthesis on polystyrene resins. These use in situ neutralization [i.e. neutralization simultaneous with coupling], high concentrations (>0.2 m) of Boc-amino acid-OBt esters plus base for rapid coupling, 100% TFA for rapid Boc group removal, and a single short (30 s) DMF flow wash between deprotection/coupling and between coupling/deprotection. Single 10 min coupling times were used throughout. Overall cycle times were 15 min for manual and 19 min for machine-assisted synthesis (75 residues per day). No racemization was detected in the _.base-catalyzed coupling step. Several side reactions were studied, and eliminated. These included: pyrrolidonecarboxylic acid formation from Gln in hot TFA-DMF; chain-termination by reaction with excess HBTU; and, chain termination by acetylation (from HOAc in commercial Boc-amino acids). The in situ neutralization protocols gave a significant increase in the efficiency of chain assembly, especially for “difficult” sequences arising from sequence-dependent peptide chain aggregation in standard (neutralization prior to coupling) Boc-chemistry SPPS protocols or in Fmoc-chemistry SPPS. Reported syntheses include HIV-1 protease(1–50,Cys.amide), HIV-1 protease(53–99), and the full length HIV-l protease(1–99). Republished with the permission of Blackwell Publishing from International Journal of Peptide Protein Research, volume 40, pp. 180–193, 1992. A preliminary account of this work was presented at the 12th American Peptide Symposium, Cambridge, MA, July 16–21, 1991 (ref. 43). Dedicated to Professor Bruce Merrifield on the occasion of his 70th birthday.     

A Novel Oxazolidine Linker for the Synthesis of Peptide Aldehydes

A reliable method for solid-phase synthesis of peptide aldehydes by using a new oxazolidine linker is described. Based on a comparative study using the usual cleavage protocol as is used for the Fmoc-based peptide synthesis, we found that this new linker is more appropriate for the synthesis of peptide aldehydes compared with the precedent acetal, semicarbazone or threonine linker. Whereas N-Acylated oxazolidines might be partially deprotected to non-N-acylated intermediates in the TFA cocktail containing several soft nucleophiles which cause significant side reactions, the new oxazolidine linker could produce the desired peptide aldehydes by simple Et₂O washing and subsequent aqueous workup in high chemical yields and purity. We demonstrate the new method is useful especially for the preparation of highly functionalized long-chain peptide aldehydes which require several scavenger chemicals in the final deprotection step. This paper is dedicated to the memory of the late Prof. R. Bruce Merrifield, who passed away May 14, 2006.     

Synthesis,antioxidative and antiviral activity of hydroxycinnamic acid amides of thiazole containing amino acid

The synthesis and the biological (antioxidant and antiviral) activities of novel hydroxycinnamic acid amides of a thiazole containing TFA.valine-4-carboxylic acid ethyl ester are reported. The amides have been synthesized from p-coumaric, ferulic and sinapic acids with the corresponding TFA.valine-thiazole-4-carboxylic acid ethyl ester using the coupling reagent N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and 4-(dimethylamino) pyridine (DMAP) as a catalyst. The antioxidant properties of the newly synthesized amides have been studied for then antioxidative activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH)* test. The newly synthesized compounds have been tested against the replication in vitro of influenza virus A (H3N2) and human herpes virus 1 and 2 (HSV-1 and HSV-2).     

Olfactory conditioning of proboscis activity in Drosophila melanogaster

Olfactory learning and memory processes in Drosophila have been well investigated with aversive conditioning, but appetitive conditioning has rarely been documented. Here, we report for the first time individual olfactory conditioning of proboscis activity in restrained Drosophila melanogaster. The protocol was adapted from those developed for proboscis extension conditioning in the honeybee Apis mellifera. After establishing a scale of small proboscis movements necessary to characterize responses to olfactory stimulation, we applied Pavlovian conditioning, with five trials consisting of paired presentation of a banana odour and a sucrose reward. Drosophila showed conditioned proboscis activity to the odour, with a twofold increase of percentage of responses after the first trial. No change occurred in flies experiencing unpaired presentations of the stimuli, confirming an associative basis for this form of olfactory learning. The adenylyl cyclase mutant rutabaga did not exhibit learning in this paradigm. This protocol generated at least a short-term memory of 15 min, but no significant associative memory was detected at 1 h. We also showed that learning performance was dependent on food motivation, by comparing flies subjected to different starvation regimes.     

Suppressing the epimerization of endothioamide peptides during Fmoc/t‐Bu‐based solid phase peptide synthesis

Despite a number of intriguing utilities associated with thioamide‐containing peptides and proteins in the context of biophysics, pharmacology and chemical biology, it has hitherto remained as one of the underexplored territories of peptidomimetics. The synthesis of long mono to multiply substituted endothioamide peptides is invariably accompanied with severe epimerization, oxoamide formation and various other undesired side reactions, resulting in messy product profiles. This has completely restrained their use as novel chemical tools for biological studies. During the chain elongation of an N‐terminally located thioamide peptide using the Fmoc/t‐Bu chemistry, it becomes vulnerable to the repetitive basic treatments as required for such chemistry. The incompatibility of thioamide moiety with bases as well as strong coupling reagents leads to epimerization as well as other side reactions due to its nucleophilicity, resulting in the loss of the stereochemical identity of the thioamidated amino acid residue. An easy‐to‐implement and efficient protocol to synthesize long (>10‐mer) endothioamide peptides, significantly suppressing epimerization and other side reactions using 10% piperidine/dimethylformamide for 1 min, is reported herein. The novelty of the protocol is shown through the efficient synthesis of a number of 10–12‐mer mono to multiply thioamide‐substituted peptides with broad substrate scopes. The utility of the protocol in the context of protein engineering and chemical protein synthesis is also shown through the synthesis of a thioamide version of the 16‐mer peptide from the B1 domain of protein G. Such a protocol to synthesize long endothioamide peptides would open up avenues toward engineering and accessing novel thiopeptide and thioprotein‐based chemical tools, the synthesis of which had been a serious hurdle thus far. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

Triacylglycerol,total fatty acid,and biomass accumulation of metabolically engineered energycane grown under field conditions confirms its potential as feedstock for drop-in fuel production

Metabolic engineering for hyperaccumulation of lipids in vegetative tissues of high biomass crops promises a step change in oil yields for the production of advanced biofuels. Energycane is the ideal feedstock for this approach due to its exceptional biomass production and persistence under marginal conditions. Here, we evaluated metabolically engineered energycane with constitutive expression of the lipogenic factors WRINKLED1 (WRI1), DIACYLGLYCEROL ACYLTRANSFERASE1 (DGAT1), and OLEOSIN1 (OLE1) for the accumulation of triacylglycerol (TAG), total fatty acid (TFA), and biomass under field conditions at the University of Florida-IFAS experiment station near Citra, Florida. TAG and TFA accumulation were highest in leaves (up to 9.9% and 12.9% of DW, respectively), followed by juice from crushed stems, stems, and roots. TAG and TFA accumulation increased up to harvest time and correlated highest with OLE1 and DGAT1 expression. Biomass dry weight, TAG, and TFA content differed greatly depending on DGAT1 and OLE1 expression in transgenic lines with similar WRI1 expression. Biomass did not significantly differ between WT and line L2 with DAGT1 and OLE1 expressed at low levels and TAG and TFA accumulating to 12- and 1.6-fold that of WT leaves, respectively. In contrast, line L13, with intron-mediated enhancement of DGAT1 expression, displayed a 245- to 330-fold increase in TAG and a 4.75- to 6.45-fold increase in TFA content compared with WT leaves and a biomass reduction of 52%. These results provide the basis for developing novel feedstocks for expanding plant lipid production and point to new prospects for advanced biofuels.     

XPairIt Docking Protocol for peptide docking and analysis

The mechanics of peptide–protein docking has long been an area of intense interest to the computational community. Here we discuss an improved docking protocol named XPairIt which uses a multitier approach, combining the PyRosetta docking software with the NAMD molecular dynamics package through a biomolecular simulation programming interface written in Python. This protocol is designed for systems where no a priori information of ligand structure (beyond sequence) or binding location is known. It provides for efficient incorporation of both ligand and target flexibility, is HPC-ready and is easily extensible for use of custom code. We apply this protocol to a set of 11 test cases drawn from benchmarking databases and from previously published studies for direct comparison with existing protocols. Strengths, weaknesses and areas of improvement are discussed.     

Survey of duckweed diversity in Lake Chao and total fatty acid,triacylglycerol, profiles of representative strains

Lemnaceae (duckweeds) are widely distributed aquatic flowering plants. Their high growth rate, starch content and suitability for bioremediation make them potential feedstock for biofuels. However, few natural duckweed resources have been investigated in China, and there is no information about total fatty acid (TFA) and triacylglycerol (TAG) composition of duckweeds from China. Here, the genetic diversity of a natural duckweed population collected from Lake Chao, China, was investigated using multilocus sequence typing (MLST). The 54 strains were categorised into four species in four genera, representing 12 distinct sequence types. Strains representing Lemna aequinoctialis and Spirodela polyrhiza were predominant. Interestingly, a surprisingly high degree of genetic diversification within L. aequinoctialis was observed. The four duckweed species revealed a uniform fatty acid composition, with three fatty acids, palmitic acid, linoleic acid and linolenic acid, accounting for more than 80% of the TFA. The TFA in biomass varied among species, ranging from 1.05% (of dry weight, DW) for L. punctata and S. polyrhiza to 1.62% for Wolffia globosa. The four duckweed species contained similar TAG contents, 0.02% mg·DW^?1. The fatty acid profiles of TAG were different from those of TFA, and also varied among the four species. The survey investigated the genetic diversity of duckweeds from Lake Chao, and provides an initial insight into TFA and TAG of four duckweed species, indicating that intraspecific and interspecific variations exist in the content and composition of both TFA and TAG in comparison with other studies.     

Preparation of an unprotected phosphotyrosine building block and its application in solid-phase synthesis of phosphopeptides

Summary A new and facile synthesis of tyrosine phosphorylated peptides has been developed.N ^α-Fmoc-Tyr(tBu)-OPfp was treated with TFA, phosphorylated with phosphorous oxychloride and the resulting phosphoric acid dichloride was hydrolysed to giveN ^α-Fmoc-Tyr(PO₃H₂)-OPfp1 in an overall yield of 98%. Compound1 was used in solid-phase peptide synthesis of phosphopeptides2, 3 and4, which are fragments of murine adipocyte lipid binding protein. The advantage of using the Pfp ester was the absence of pyrophosphates and other byproducts.     

黄芪总黄酮对DNA损伤防护作用的研究

用DNA解旋荧光检测法(FADU)研究了黄芪总黄酮(TFA)对γ射线和H₂O₂所致V79细胞DNA链断裂的防护作用. 结果表明TFA对这两种损伤因子所致的DNA损伤均有不同程度的防护作用, 当TFA浓度达到0.4g/L和0.6g/L时, 分别对H₂O₂和γ射线所致的损伤有保护作用(P<0.05), 而浓度增至0.8g/L和1.2g/L时, 分别对两种因素所致的DNA链断裂损伤有非常显著的防护效果(P<0.01), 对H₂O₂的防护效果优于对γ射线.     

Facile preparation of the N‐acetyl‐glucosaminylated asparagine derivative with TFA‐sensitive protecting groups useful for solid‐phase glycopeptide synthesis

In this study, a novel N‐acetyl‐glucosaminylated asparagine derivative was developed. This derivative carried TFA‐sensitive protecting groups and was derived from commercially available compounds only in three steps. It was applicable to the ordinary 9‐fluorenylmethoxycarbonyl (Fmoc)‐based solid‐phase peptide synthesis (SPPS) method, and the protecting groups on the carbohydrate moiety could be removed by a single step of TFA cocktail treatment generally used for the final deprotection step in Fmoc‐SPPS. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.     

Conformational aspect of polypeptide structure. XLIV. Conformational transitions of poly(N-methyl-alanines) induced by trifluoroacetic acid

The synthesis of poly(N-methyl-L -alanine) and poly (N-methyl-DL -alanine) are described. The polymers were examined by 220 MHz high-resolution nuclear magnetic resonance (nmr) and circular dichroism (CD). The results demonstrate that poly(N-methyl-L -alanine) exists as an ordered helical structure with all the amide bonds in the trans configuration in appropriate solvents. As trifluoroacetic acid (TFA) is added to the solutions of the polymer in helix-supporting solvents, resonances corresponding to both trans and cis amide conformations of N-methyl, C-methyl, and α-CH are observed. The presence of both the trans and the cis peptide bonds in a polymer chain disrupts the ordered structures. Our conclusions from CD data are in agreement with the nmr results. Ultracentrifugation shows that degradation of the polymer chain does not occur during the TFA treatment.     

Differences among captive callitrichids in the digestive responses to dietary gum

Although many members of the Callitrichidae, a monophyletic family of small, New World monkeys, have been observed to feed on plant exudates, available field data support the generalization that pygmy and common marmosets (Cebuella pygmaea and Callithrix jacchus) feed on gums to a greater extent than most other callitrichids. Because microbial fermentation is required for vertebrates to digest gums, gum-feeding primates may react differently to dietary gum from their relatives that eat little gum. To test this hypothesis, digestion trials were conducted on animals from the two marmoset species, two tamarin species (Saguinus fuscicollis and S. oedipus), and a species of lion tamarin (Leontopithecus rosalia). These species span the range of body sizes within the Callitrichidae. All animals were fed two variations of a homogeneous diet, which differed only in that gum arabic was added to one. Transit time of digesta (TFA) and digestive efficiency (as measured by the coefficients of apparent digestibility of dry matter and energy [ADDM and ADE, respectively]) were compared between diets for each individual. As predicted, the digestive responses of marmosets differed from the responses of the other study species. In marmosets, TFA tended to be longer when gum was added to the diet, while TFA did not change in the other three species. Digestive efficiency decreased in tamarins and lion tamarins with the addition of gum to the diet; marmoset digestive efficiency was unaffected by diet. The results of this research are consistent with the hypothesis that marmosets have digestive adaptations that aid in the digestion of gum that other callitrichids lack. © 1996 Wiley-Liss, Inc.     

Parallel synthesis of peptide libraries using microwave irradiation

The application of microwave irradiation to solid-phase peptide synthesis increases product purity and reduces reaction time. Parallel synthesis in 96-well polypropylene filter plates with microwave irradiation is an efficient method for the rapid generation of combinatorial peptide libraries in sufficient purity to assay the products directly for biological activity without HPLC purification. In this protocol, the solid-phase support is arrayed into each well of a 96-well plate, reagents are delivered using a multichannel pipette and a microwave reactor is used to complete peptide coupling reactions in 6 min and Fmoc-removal reactions in 4 min under temperature-controlled conditions. The microwave-assisted parallel peptide synthesis protocol has been used to generate a library of difficult hexa-beta-peptides in 61% average initial purity (50% yield) and has been applied to the preparation of longer alpha- and beta-peptides. Using this protocol, a library of 96 different hexapeptides can be synthesized in 24 h (excluding characterization).