A hydrazide

A special problem arises in the preparation of secondary amines. These compounds are highly nucleophilic, and alkylation of an amine with alkyl halides cannot be expected to stop at any specifle stage. Secondary amides, however, can be monoalkylated and lydrolyzed or be reduced to secondary amines (p. 11 If.). In the elegant synthesis of phenyl- phrine an intermediate -hydroxy isocyanate (from a hydrazide and nitrous acid) cyclizes to pve an oxazolidinone which is monomethylated. Treatment with strong acid cleaves the cyclic irethan. 

Reaction of a hydrazide (128) with phosgeneiminium chloride (115) led to the 2-dimethylamino-l,3,4-oxadiazole (129) in 90% yield (75AG(E)806). The 1,3,4-thiadiazole system was also obtained in an analogous reaction in which the dithioimidate (130) underwent reaction with the thiohydrazide (131). Depending on the nature of X in (131), the 2-substituent in the resultant 1,3,4-thiadiazole (132) may be varied (80ZC413). Although (130)... 

H2NNH2, MeOH, reflux, 60 min, 100% yield.In this case the ester is converted to a hydrazide. 

In the patent literature, Aventis has published a patent application covering hydroxamic acid containing macrocycles (39) with the same general template as the A-formyl-A-hydroxylamine macrocycles described above [114] and GlaxoSmithKline has published an application on macrocyclic PDF inhibitors containing a hydrazide scaffold (40) [115], No data has been published on these inhibitors to date. 

Hurwitz et al. [256] used a hydrazide derivative of poly(glutamic acid) as a carrier for daunamycin. This was less toxic than free drug against mouse lymphoma in vitro, but it was as effective, or more effective, against the same lymphoma in vivo. 

The formation of compounds 2-503 proceeds via the addition of a hydrazide 2-500 onto the central carbon of an isothiocyanate 2-499. Subsequent cyclization by attack of the hydrazide nitrogen in the formed 2-501 onto the nitrile moiety gives intermediate 2-502. A further attack of the newly formed amidine nitrogen onto the carbonyl group followed by extrusion of water affords the triazoloquinolines 2-503. 

Polysaccharides, glycoproteins, and other glycoconjugates therefore may be specifically labeled on their carbohydrate portions by creating aldehyde functionalities and subsequently derivatizing them with another molecule containing an amine or a hydrazide group. This route of derivatization is probably the most common way of modifying carbohydrates. 

Figure 1.70 AMBH is a hydrazide-containing compound that reacts with carbonyl groups to form hydrazone bonds. The free thiol can be used for subsequent conjugation reactions.

Figure 1.107 The N-terminal aldehyde group on a peptide formed from periodate oxidation of serine or threonine residues can be conjugated with a hydrazide-containing molecule to produce a hydrazone bond.

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.

In aqueous solutions, the easiest method for forming this type of bond is to use the water-soluble carbodiimide EDC (Chapter 3, Section 1.1). For proteins and other water-soluble macromolecules, EDC reacts with their available carboxylate groups to form an intermediate, highly reactive, o-acylisourea. This active ester species may further react with nucleophiles such as a hydrazide to yield a stable imide product (Figure 1.109). 

Derivatives of hydrazine, especially the hydrazide compounds formed from carboxylate groups, can react specifically with aldehyde or ketone functional groups in target molecules. Reaction with either group creates a hydrazone linkage (Reaction 44)—a type of Schiff base. This bond is relatively stable if it is formed with a ketone, but somewhat labile if the reaction is with an aldehyde group. However, the reaction rate of hydrazine derivatives with aldehydes typically is faster than the rate with ketones. Hydrazone formation with aldehydes, however, results in much more stable bonds than the easily reversible Schiff base interaction of an amine with an aldehyde. To further stabilize the bond between a hydrazide and an aldehyde, the hydrazone may be reacted with sodium cyanoborohydride to reduce the double bond and form a secure covalent linkage. 

Bis-hydrazide-containing molecules also can be used to activate soluble polymeric sub-stances-containing aldehyde groups. For instance, dextran may be periodate oxidized to create numerous formyl functionalities on each molecule. Subsequent reaction with a homobifunctional hydrazide in large excess results in a hydrazide-activated polymer having multivalent-binding capability toward aldehydes or ketones (Chapter 25, Section 2.2). Insoluble support matrices suitable for affinity chromatography have been activated in a similar fashion to create the hydrazide derivative (O Shannessy and Wilchek, 1990). 

Figure 4.30 Carbohydrazide can be used to transform an aldehyde residue into a hydrazide group.

The carbonyl-reactive group on these crosslinkers is a hydrazide that can form hydrazone bonds with aldehyde residues. To utilize this functional group with carbohydrate-containing molecules, the sugars first must be mildly oxidized to contain aldehyde groups by treatment with sodium periodate. Oxidation with this compound will cleave adjacent carbon-carbon bonds which possess hydroxyl groups, as are abundant in polysaccharide molecules (Chapter 1, Sections 2 and 4.4). 

Lissamine rhodamine B sulfonyl hydrazine is a hydrazide derivative of sulforhodamine B that can spontaneously react with aldehyde- or ketone-containing molecules to form a covalent,... 

Figure 9.48 A cyanine dye containing a hydrazide group can be used to label glycans at their reducing end or other reducing sugars, forming a hydrazone linkage. Glycoproteins also can be labeled after periodate oxidation to form aldehyde groups.

BNAH may be coupled to reducing sugars without reduction, since the linkage formed between a hydrazide and an aldehyde is much more stable than that with an amine. 

In some cases, the ability to modify glycans at the reducing end without reduction preserves the carbohydrate s native structure sufficiently to allow interactions with proteins that would otherwise not interact if the bond were reduced. Therefore, depending on the ultimate use of the biotinylated carbohydrate, using a hydrazide mediated conjugation process can have advantages over the use of amine-biotin compounds. 

Figure 11.21 BNAH contains a hydrazide group that can be used to label the reducing end of released glycans through the formation of a hydrazone bond.

Another route to the formation of a hydrazide on a surface is to use an aldehyde-containing particle (such as HEMA/acrolein copolymers) and subsequently modify the aldehydes to form hydrazone linkages with bis-hydrazide compounds, which then can be stabilized by reduction with sodium cyanoborohydride (Chapter 2, Section 5). The resulting derivative contains terminal hydrazides for immobilization of carbonyl ligands (see Figure 14.18). 

Figure 14.18 Carboxylate-particles or aldehyde-particles can be modified with the carbohydrazide in excess to create a hydrazide-particle that can be used to couple with aldehyde-containing molecules.

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