Adipic acid hydrazide

A keto acid ester such as allyl levulinate may be crosaUxlced by adipic acid hydrazide, and some unsaturated diketones or keto-acids would probably react similarly. Polymers have been descriled in bead form, but conditions could probably be found to enstire emulsion stability. (55)... 

Putrescine dihydrochloride has been prepared by the Hofmann degradation of adipamide 3.. s by the Curtius degradation of adipyl hydrazide through the urethane by the Curtius degradation of adipyl azide obtained from adipyl chloride and sodium azide by the Schmidt degradation of adipic acid with hydrogen azide by the reduction of succinonitrile, succinaldoxime, or 7-phthalimidobutyronitrile with sodium and from N-ben-zoyl-7-iodobutylamine ... 

Aldehyde-containing macromolecules will react spontaneously with hydrazide compounds to form hydrazone linkages. The hydrazone bond is a form of Schiff base that is more stable than the Schiff base formed from the interaction of an aldehyde and an amine. The hydrazone, however, may be reduced and further stabilized by the same reductants utilized for reductive amination purposes (Chapter 3, Section 4.8). The addition of sodium cyanoborohydride to a hydrazide-aldehyde reaction drives the equilibrium toward formation of a stable covalent complex. Mallia (1992) found that adipic acid dihydrazide derivatization of periodate-oxidized dextran (containing multiple formyl functionalities) proceeds with much greater yield when sodium cyanoborohydride is present. 

Figure 1.108 Glycoproteins that have been treated with sodium periodate to produce aldehyde groups can be further modified with adipic acid dihydrazide to result in a hydrazide derivative.

Figure 1.109 Carboxylate groups on proteins may be modified with adipic acid dihydrazide in the presence of a carbodiimide to produce hydrazide derivatives.

Carboxylic acids may be covalently modified with adipic acid dihydrazide or carbohydrazide to yield stable imide bonds with extending terminal hydrazide groups. Hydrazide functionalities don t spontaneously react with carboxylate groups the way they do with aldehyde groups (Section 4.5, this chapter). In this case, the carboxylic acid first must be activated with another compound that makes it reactive toward nucleophiles. In organic solutions, this may be accomplished by using a water-insoluble carbodiimide (Chapter 3, Section 1.4) or by creating an intermediate active ester, such as an NHS ester (Chapter 2, Section 1.4). 

Most proteins contain an abundance of carboxylic acid groups from C-terminal functionalities and aspartic and glutamic acid side chains. These groups are readily modified with bis-hydrazide compounds to yield useful hydrazide-activated derivatives. Both carbohydrazide and adipic acid dihydrazide have been employed in forming these modifications using the carbodi-imide reaction (Wilchek and Bayer, 1987). 

Add to the tube 7.5 pi of RNA or DNA containing a 5 -phosphate group. The concentration of the oligonucleotide should be 7.5-15 nmol or total of about 57-115.5 pg. Also, immediately add 5 pi of 0.25 M adipic acid dihydrazide or carbohydrazide dissolved in 0.1 M imidazole, pH 6.0. Because EDC is labile in aqueous solutions, the addition of the oligo and fezs-hydrazide/imidazole solutions should occur quickly. 

Figure 23.9 Reaction of adipic acid dihydrazide with (strept)avidin produces a hydrazide derivative that is highly reactive toward periodate-oxidized polysaccharides.

To make an amine derivative of dextran, dissolve ethylene diamine (or another suitable diamine) in 0.1 M sodium phosphate, 0.15 M NaCl, pH 7.2, at a concentration of 3 M. Note Use of the hydrochloride form of ethylene diamine is more convenient, since it avoids having to adjust the pH of the highly alkaline free-base form of the molecule. Alternatively, to prepare a hydrazide-dextran derivative, dissolve adipic acid dihydrazide (Chapter 4, Section 8.1) in the coupling buffer at a concentration of 30 mg/ml (heating under a hot water tap may be necessary to completely dissolve the hydrazide compound). Adjust the pH to 7.2 with HC1 and cool to room temperature. 

Hydrazide groups can react with carbonyl groups to form stable hydrazone linkages. Derivatives of proteins formed from the reaction of their carboxylate side chains with adipic acid dihydrazide (Chapter 4, Section 8.1) and the water-soluble carbodiimide EDC (Chapter 3, Section 1.1) create activated proteins that can covalently bind to formyl residues. Hydrazide-modified enzymes prepared in this manner can bind specifically to aldehyde groups formed by mild periodate oxidation of carbohydrates (Chapter 1, Section 4.4). These reagents can be used in assay systems to detect or measure glycoproteins in cells, tissue sections, or blots (Gershoni et al., 1985). 

The activation of enzymes using adipic acid dihydrazide and EDC is identical to the procedure outlined for the modification of (strept)avidin (Chapter 23, Section 5). Alternatively, hydrazide groups may be created on enzymes using the heterobifunctional chemoselective reagents described in Chapter 17, Section 2. 

The following protocol describes the modification of DNA or RNA probes at their 5 -phosphate ends with a bis-hydrazide compound, such as adipic acid dihydrazide or carbohydrazide. A similar procedure for coupling the diamine compound cystamine can be found in Section 2.2 (this chapter). 

Adipic Acid Atlde or Adipylazide (Adipinyl-azide), N3. CO(CH,)4CO. OH-not found in Beil or CA through 1956 Adipic Acid Diazide or Adipyldiazide (Adi piny Idi azide), Ns, CO(CH()4CO. N mw 196.17,N 42.84% col oil solidifying at -1° to wh crysts bp-expl viol on heating directly or under w easily sol in ale or eth. Was first prepd in 1915 from adipyl hydrazide,... 

The reaction of an excess of adipic acid dihydrazide with aldehyde groups present on proteins or other molecules will result in modified proteins containing alkyl-hydrazide groups (Fig. 96). Another bis-hydrazide compound, carbohydrazide, also may be employed with similar results, except that the spacer afforded through its use is considerably shorter. Target aldehydes may be created on macromolecules according... 

Adenosine triphosphate, when oxidized to its dialdehyde with periodate and condensed with Sepharose-adipic (or sebacic) acid hydrazide, has been used to isolate poly(A) polymerase, and also myosin and its fragments from different sources. A poly(A)-linked resin has been prepared by condensing 4,4 -diaminodiphenyl-methane with periodate-oxidized starch, reducing the resulting Schiff bases with borohydride, diazotizing, and then coupling with the polynucleotide. The resulting material may be used to isolate poly(U) sequences. 

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