Peptide synthesizer

In synthetic target molecules esters, lactones, amides, and lactams are the most common carboxylic acid derivatives. In order to synthesize them from carboxylic acids one has generally to produce an activated acid derivative, and an enormous variety of activating reagents is known, mostly developed for peptide syntheses (M. Bodanszky, 1976). In actual syntheses of complex esters and amides, however, only a small selection of these remedies is used, and we shall mention only generally applicable methods. The classic means of activating carboxyl groups arc the acyl azide method of Curtius and the acyl chloride method of Emil Fischer. 

In peptide syntheses, where partial racemization of the chiral a-carbon centers is a serious problem, the application of 1-hydroxy-1 H-benzotriazole ( HBT") and DCC has been very successful in increasing yields and decreasing racemization (W. Kdnig, 1970 G.C. Windridge, 1971 H.R. Bosshard, 1973), l-(Acyloxy)-lif-benzotriazoles or l-acyl-17f-benzo-triazole 3-oxides are formed as reactive intermediates. If carboxylic or phosphoric esters are to be formed from the acids and alcohols using DCC, 4-(pyrrolidin-l -yl)pyridine ( PPY A. Hassner, 1978 K.M. Patel, 1979) and HBT are efficient catalysts even with tert-alkyl, choles-teryl, aryl, and other unreactive alcohols as well as with highly bulky or labile acids. 

The cyclopeptide described above was tailored to form stable potassium complexes. It is one of the very few examples of complex peptide syntheses which do not lead to a natural compound. 

Memfield successfully automated all the steps m solid phase peptide synthesis and computer controlled equipment is now commercially available to perform this synthesis Using an early version of his peptide synthesizer m collaboration with coworker Bemd Gutte Memfield reported the synthesis of the enzyme ribonuclease m 1969 It took them only SIX weeks to perform the 369 reactions and 11 391 steps necessary to assemble the sequence of 124 ammo acids of ribonuclease... 

Urodilantin, a 32-amino acid natriuretic peptide synthesized by the kidney and found in urine but not in plasma, is beheved to compHcate the interpretation of the natriuretic effects of ANP (58). There have been reports of natriuretic peptides related to ANP that have been isolated from several sources, including the brain, eg, brain natriuretic peptide (BNP) [114471-18-0] (59—61). 

One widely used method of formation of protected compounds involves polymer-supported reagents, with the advantage of simple workup by filtration and automated syntheses, especially of polypeptides. Polymer-supported reagents are used to protect a terminal — COOH group as a polymer-bound ester (RCOOR —( ) during peptide syntheses, to protect primary alcohols as... 

Hydrazides have been used in penicillin and peptide syntheses. In the latter syntheses they are converted by nitrous acid to azides to facilitate coupling. 

Two disadvantages are associated with the use of S-acetyl or 5-benzoyl derivatives in peptide syntheses (a) base-catalyzed hydrolysis of 5-acetyl- and 5-benzoylcys-teine occurs with /S-elimination to give olefinic side products, CH2=C-(NHPG)CO—(b) the yields of peptides formed by coupling an unprotected amino group in an 5-acylcysteine are low because of prior S-N acyl migration. ... 

A number of 5-sulfenylthiocarbonates have been prepared to protect thiols. A benzyl derivative, R =CH2Ph, is stable to trifluoroacetic acid (25°, 1 h) and provides satisfactory protection during peptide syntheses a r-butyl derivative, R = r-Bu, is too labile in base to provide protection. A methyl derivative, R =CH3, has... 

A -Nitroso derivatives, prepared from secondary amines and nitrous acid, are cleaved by reduction (H2/Raney Ni, EtOH, 28°, 3.5 h CuCl/concd. HCl"). Since many V-nitroso compounds are carcinogens, and because some racemization and cyclodehydration of V-nitroso derivatives of V-alkyl amino acids occur during peptide syntheses, V-nitroso derivatives are of limited value as protective groups. 

The procedure described is essentially that of Shioiri and Yamada. Diphenyl phosphorazidate is a useful and versatile reagent in organic synthesis. It has been used for racemlzatlon-free peptide syntheses, thiol ester synthesis, a modified Curtius reaction, an esterification of a-substituted carboxylic acld, formation of diketoplperazines, alkyl azide synthesis, phosphorylation of alcohols and amines,and polymerization of amino acids and peptides. - Furthermore, diphenyl phosphorazidate acts as a nitrene source and as a 1,3-dipole.An example in the ring contraction of cyclic ketones to form cycloalkanecarboxylic acids is presented in the next procedure, this volume. 

The formation of vinylogous amides from primary amines and -dicarbonyl compounds gives rise to hydrolyzable amine derivatives with greatly decreased nucleophilicity of the nitrogen function. Thus these derivatives have found some use as protecting groups in peptide syntheses 617-619). 

Catalytic hydrogenolysis of an O-benzyl protective group is a mild, selective method introduced by Bergmann and Zervas to cleave a benzyl carbamate (>NC0-0CH2C6H5 —> >NH) prepared to protect an amino group during peptide syntheses. The method has also been used to cleave alkyl benzyl ethers, stable compounds prepared to protect alkyl alcohols benzyl esters are cleaved by catalytic hydrogenolysis under neutral conditions. 

It has been used in peptide syntheses, but is very labile to acids and bases. ... 

Robotic peptide synthesizers are now used to automatically repeat the coupling, washing, and deprotection steps with different amino acids. Each step occurs in high yield, and mechanical losses are minimized because the peptide intermediates are never removed from the insoluble polymer until the final step. Using this procedure, up to 25 to 30 mg of a peptide with 20 amino acids can be routinely prepared. 

DNA synthesizers operate on a principle similar to that of the Merrifield solid-phase peptide synthesizer (Section 26.8). In essence, a protected nucleotide is covalently bonded to a solid support, and one nucleotide at a time is added to the growing chain by the use of a coupling reagent. After the final nucleotide has been added, all the protecting groups are removed and the synthetic DNA is cleaved from the solid support. Five steps are needed ... 

The mechanistic analogy to the Streckcr synthesis becomes obvious in the addition of the isocyanide to the imine to produce the a-amino nitrilium intermediate. Since all four components are involved in this step, it might be expected that every chiral component (chiral groups R1, R2, R3, R4) contributes to diastereofacial differentiation in the nucleophilic attack on the imine. However, in peptide syntheses by four-component condensation5, the chiral isocyanide or a chiral carboxylic acid component has only limited influence on the diastereoselectivity of the a-amino amide formation5. 

A six aminoacid neurotensin-like peptides synthesized in the same propeptide precursor as neurotensin. 

PEPTIDE SYNTHESES USING N-ETHYL-5-PHENYLIS0XAZ0LIUM-3 -SULF0NATE CARBOBENZOXY-l-ASPARAGINYL-l-LEUCINE METHYL ESTER AND JV-CARBOBENZOXY-3-HYDROXY-l-PROLYLGLYCYLGLYCINE ETHYL ESTER... 

Fig. 10 Liberty (CEM Corporation), the first automated microwave solid-phase peptide synthesizer, is shown on the left-, the available reaction vessel setup for solid-phase synthesis is depicted on the right...

At this time, a solid phase automatic peptide synthesizer was acquired, so we attempted total synthesis of Sh I. A solid phase synthesis of Ax I (antho-pleurin A) had previously been reported in an abstract (36). The synthetic Ax I possessed only 11% of the toxicity of the natural toxin. 

The first peptide syntheses were also initiated in Bonn. In the later years of his research activity, Helferich concentrated particularly on the chemistry of the sulfonamides. The chemistry of the sultams was of particular interest to him. As a spin-off from this work he developed a psychotropic drug, namely, Ospolot. 

Drivers for Performing Peptide Syntheses in Micro Reactors... 

Beneficial Micro Reactor Properties for Peptide Syntheses... 

Atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP) are members of a family of so-called natriuretic peptides, synthesized predominantly in the cardiac atrium, ventricle, and vascular endothelial cells, respectively (G13, Y2). ANP is a 28-amino-acid polypeptide hormone released into the circulation in response to atrial stretch (L3). ANP acts (Fig. 8) on the kidney to increase sodium excretion and glomerular filtration rate (GFR), to antagonize renal vasoconstriction, and to inhibit renin secretion (Ml). In the cardiovascular system, ANP antagonizes vasoconstriction and shifts fluid from the intravascular to the interstitial compartment (G14). In the adrenal cortex, ANP is a powerful inhibitor of aldosterone synthesis (E6, N3). At the hypothalamic level, ANP inhibits vasopressin secretion (S3). It has been shown that some of the effects of ANP are mediated via a newly discovered hormone, called adreno-medullin, controlling fluid and electrolyte homeostasis (S8). The diuretic and blood pressure-lowering effect of ANP may be partially due to adrenomedullin (V5). 

Nitrophenyl esters of amino acids, which are important for peptide syntheses, have been obtained in a one-pot reaction from TV-protected amino acids, CDI, and /j-nitro-phenol at room temperature however, better yields of these esters could be achieved by use of TV-trifluoroacetylimidazole. In this reaction a mixed anhydride is presumably formed as an intermediate, which then acylates the alcohol component [17]... 

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