Selective Hydrolysis Methods

Determination of C-terminal amino acid residues by use of hydrazine. 

CHAPTERS Protein Isolation and Determination of Amino Acid Sequence 

Cleavage of a disulfide bond (cystine residue) by performic acid. 

Hydrolysis of Polypeptides at Specific Sites by Selective Reagents 

Amino acid Amino acid residue 1 residue 2 (carboxyl side) (amino side) 

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Another way of applying the selective extraction method directly on the initial solution is to produce a solution of low acidity. This can be achieved by using the hydrofluoride method for fluorination and decomposition of raw material. As was discussed in Paragraph 8.2.2, the raw material is fluorinated by molten ammonium hydrofluoride yielding soluble complex fluorides of ammonium and tantalum or niobium. The cake obtained following fluorination is dissolved in water, leading to a solution of low initial acidity that is related for the most part to the partial hydrolysis of complex fluoride compounds. The acidity of the solution is first adjusted to ensure selective tantalum extraction. In the second step, the acidity of the raffinate is increased to provide the necessary conditions for niobium extraction. 

Hydroxydeethoxylation of the 2>H- -benzazepine 3 in refluxing methanolic potassium hydroxide (Method A) is accompanied by selective hydrolysis and loss of the 3-carboxylate group.20 Hydrolysis in concentrated hydrochloric acid, however, is rapid and yields only the bcnz-azepinone 4 (Method B). 

It was reported that III could be separated from IV by selective hydrolysis of IV (10), but these results were later disputed (12). Indeed, in our hands purification of III by selective hydrolysis of IV to isophorone using the literature method (10) failed. There was no observed change In the product mixture. 

A suitable method for this was introduced by Caspersson (mid-1920s-ca. 1940) who designed and successfully exploited UV microscopy so that the extent of the absorbtion could be determined quantitatively. Selective hydrolysis by RNAase or DNAase was used so that the DNA content of the cell could be estimated. The procedure still required reproducible preparation of sections to allow light to be transmitted and the enzymes to get access to the nucleic acids. 

Semi-synthetic penicillins are accessed from 6-aminopenicillanic acid, (6-APA), derived from fermented penicillin G. Starting materials for semi-synthetic cephalosporins are either 7-aminodesacetoxycephalosporanic acid (7-ADCA), which is also derived from penicillin G or 7-aminocephalosporanic acid (7-ACA), derived from fermented cephalosporin C (Scheme 1.10). These three key building blocks are produced in thousands of tonnes annually worldwide. The relatively labile nature of these molecules has encouraged the development of mild biocatalytic methods for selective hydrolysis and attachment of side chains. 

The results of selective hydrolysis together with the percentages of isomers formed suggest that the facility for hydrolytic cleavage is in the order C-10 > C-8 > C-6. This order agrees with the order of bond energies between bromine and carbon atoms as calculated by the MINDO/3 method for the five isomeric monobromobenzo[6]tropoxazines. 

The problem of selective hydrolysis of the acetonlde was studied next. The most practical method consisted of a treatment with acetic acid In aqueous tetrahydrofuran at 65°C which gave a mixture of dlol 103 (major) and trlol 104 (minor) In 77 and 15% yields respectively. Trlol 104 could be converted to 103 by a sequence Involving formation of a 2 3 -orthoester, conversion to the corresponding dl-BOM derivative, then mild hydrolysis. 

Partial hydrolysis of a fully methylated polysaccharide and investigation of the products as just described may provide valuable information, even when the partial hydrolysis is not very selective. The method has been used in studies of the Klebsiella type 38 (Ref. 17) and type 52 (Ref. 28) capsular polysaccharides. In their studies of the Klebsiella type 56 capsular polysaccharide, Choy and Dutton used a modification of this method.29... 

The preparation of -substituted derivatives is more difficult and different methods have been used in the various series. 2-Tritylpyrrole undergoes electrophilic substitution selectively at the 3-position (Section 3.3.1.4.3) and the trityl group can then be removed. Again, in the pyrrole series the selective hydrolysis of a-C02Et in alkali and of (3-C02Et in acid, followed by decarboxylation, allows the introduction of (3-substituents into compounds such as 2,4-dialkylpyrrole-3,5-dicarboxylic esters to afford 3-substituted 2,4-dialkylpyrroles (Section 33.3.3.1). 

Enantioselective enzymatic amide hydrolyses can also be applied for the preparation of optically active organosilicon compounds. The first example of this is the kinetic resolution of the racemic [l-(phenylacetamido)ethyl] silane rac-84 using immobilized penicillin G acylase (PGA E.C. 3.5.1.11) from Escherichia coli as the biocatalyst (Scheme 18)69. (R)-selective hydrolysis of rac-84 yielded the corresponding (l-aminoethyl)silane (R)-85 which was obtained on a preparative scale in 40% yield (relative to rac-84). The enantiomeric purity of the biotransformation product was 92% ee. This method has not yet been used for the synthesis of optically active silicon compounds with the silicon atom as the center of chirality. 

Both linear and supercoiled double-stranded DNA can be used as substrate for site-selective scission. Typical methods for the scission of linear double-stranded DNA are given in Sections 7.6.2.1 and 7.6.2.2. The substrate is obtained by treating supercoiled PBR322 plasmid DNA by a restriction enzyme EcoRI. Here, the plasmid DNA is cut at one-site, providing a linear DNA (this DNA is known as form III). Section 7.6.2.3 describes the method for site-selective hydrolysis of supercoiled DNA. Figure 7.9(a)... 

In the next example, a lithium base (lithium diethylamide) is used to form the aza-enolate. The ease of imine cleavage in acid is demonstrated by the selective hydrolysis to the aldehyde without any effect on the acetal introduced by the alkylation step. The product is a mono-protected dialdehyde— difficult to prepare by other methods. 

This method is based on the preferential oxidation of the disordered regions by sodium metaperiodate [266,267], Conditions are selected so that the reaction is confined as far as possible to the Malaprade course resulting in the formation of 2,3-dialdehyde units. The course of the reaction is followed by measuring the oxidant consumption from the amount of periodate consumed. From plots of log oxidant consumption against time, a measure of the fraction of ordered material can be calculated analogous to that of the acid hydrolysis method. 

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