It is well established that many of the substrates of intermediary metabolism undergo enzymatic reaction only in the form of phosphate esters, even though the reaction may take place at a site removed from the phosphate group. For example, the glucose molecule is phosphorylated as it enters the living cell, and, in its subsequent transformation through fructose and the trioses, phosphate ester groups are always present. Although it has been shown that the contribution of the 6-phosphate group in fructose diphosphate is to prevent pyranose ring-formation (132), it would appear possible that in many instances the function of the phosphate group is to bind the substrate to the enzyme molecule. If so, the question arises whether a similar chemical structure, such as a phosphonic acid group, could carry out the same function.

Accordingly, the synthesis was undertaken of a number of phosphonate analogues of phosphorylated intermediates of glycolysis, in particular the triose phosphates. During the course of these preparative studies, the emphasis shifted from the original biochemical consideration to the novel and interesting chemistry encountered in the preparation of phosphonic acids possessing ketone and hydroxyl groups in the β- or γ-positions.

Synthesis Of Phosphonates From Alcohols

The most direct approach to the synthesis of phosphonate analogues of phosphate esters involves the replacement of a hydroxyl group with a phosphonate group. Heretofore, the only general procedure for this transformation was the replacement of the hydroxyl group first by a halogen atom (X = Cl, Br, I) followed by treatment of the alkyl halide with either a trialkyl phosphite (Michaelis-Arbuzov reaction, eq. 1, Fig. 39) or a sodium dialkyl phosphonate (Nylen reaction, eq. 2). The replacement of hydroxyl by halogen often involves reaction conditions not well tolerated by many sensitive physiological substances. An alternate method has been developed (eq. 3) in which the hydroxyl group is converted under mild conditions into its p-toluenesulfonate or methanesulfonate ester, which, on treatment with a sodium dialkyl phosphonate, yields the desired phosphonate ester (143). Acid hydrolysis of the latter furnishes the free phosphonic acid. Alkyl tosylates were found to be intermediate between alkyl chlorides and bromides in their reactivity toward sodium diethyl phosphonate (27). During the course of these studies an improved method for the preparation of sodium diethyl phosphonate was discovered (27).

Fig. 39.-Organophosphorus reactions.

Fig 39.-Continued.

β-Ketophosphonate And Enol Phosphate Esters

It has long been recognized that chlorine or bromine substituents adjacent to a carbonyl group do not react normally with either trialkyl phosphites or sodium dialkyl phosphonates. More recently, it has been shown (Per-kow, W. Ber. 87:755, 1954) that the reaction products actually are enol phosphate rather than β-ketophosphonate esters (eq. 4). The abnormal reaction of a-haloketones is considered to arise from the polarization of the carbonyl group by the electronegative chlorine or bromine atom. Therefore it appeared reasonable that an a-iodoketone, in which the halogen not only is less electronegative but also more reactive in simple nucleophilic displacement, would be more likely to undergo a normal Michaelis-Arbuzov reaction. Accordingly, the reaction of iodoacetone with triethyl phosphite was investigated (no) and found to yield the first example of the hitherto unknown β-ketophosphonate esters (eq. 5).

An alternate route to the same β-ketophosphonate ester was provided by the treatment of a.a'-dichloracetone with two moles of triethyl phosphite (no). This reaction yields a mixed phosphonate-enol phosphate (eq. 6), which, on acid catalyzed ethanolysis of the enol phosphate linkage, gives the /3-ketophosphonate. On treatment of the phosphonate-enol phosphate with base, the elements of diethyl hydrogen phosphate are eliminated to form an allene which, under most conditions, is converted into an alkynyl-phosphonate (eq. 6a). An extensive study of a variety of enol phosphates (d5) showed that the ready elimination of dialkyl phosphate upon treatment with base is a property only of enol phosphate esters which contain an allylic hydrogen atom activated either by a phosphonate or a carboxylate group. It was further observed that hydrogenation of simple enol phosphate esters over palladium yields trialkyl phosphates, but over platinum reductive cleavage takes place to yield dialkyl hydrogen phosphates (no).

γ-Ketophosphonate Esters And Acids

A simple and efficient synthesis of γ-ketophosphonates (141) is afforded by the smooth reaction between triethyl phosphite and either the methiodide or hydrochloride of the Mannich base, obtained by treatment of a methyl ketone with formaldehyde and a secondary amine (eq. 7). Acid hydrolysis of the phosphonate ester furnishes the free 7-ketophosphonic acid.

β-Hydroxy- And β,γ-Dihydroxyphosphonates

In the attempt to prepare hydroxylated propylphosphonic acids and esters, it was found that chlorine and bromine atoms adjacent to a hydroxy sub-stituent do not react normally with either triethyl phosphite or sodium diethyl phosphonate to furnish β-hydroxyphosphonates. The reaction of propylene oxide with sodium diethyl phosphonate rather unexpectedly gave tetraethyl propyl-1,2-diphosphonate. A convenient route (152) to β-hydroxyphosphonates was afforded by the peroxide-catalyzed chain reaction of diethyl phosphonate with an enol acetate to yield a β-acctoxy-phosphonate from which the acetyl group can be removed selectively by treatment with alcoholic barium hydroxide (eq. 8). Treatment of diethyl β-acetoxypropylphosphonate with hydrochloric acid gave free β-hy-droxypropylphosphonic acid, a hygroscopic liquid which could be isolated only as its crystalline cyclohexylamine salt.

β,γ-Dihydroxypropylphosphonic acid (152) was obtained as its barium salt by the hydrolysis of diethyl β,γ-epoxypropylphosphonate which was obtained by the treatment of epibromohydrin with triethyl phosphite (eq- 9).

Reaction Of Organic Disulfides With Trivalent Phosphorus Reagents

In analogy to their reactions with alkyl halides, alkyl tosylates, and Mannich base salts, trialkyl phosphites (25, 112, d7) and sodium dialkyl phos-phonates (24, d7) were found to react with organic disulfides to produce O.O.S-trialkyl phosphorothioates. This reaction involves heterolytic cleavage of the disulfide linkage by the nucleophilic phosphorus atom to displace a mercaptide anion. With unsymmetrical disulfides, cleavage takes place exclusively in one direction so as to displace the more stable of the two possible mercaptide anions.

In the reaction of trialkyl phosphites (eq. 10) the displaced mercaptide ion appears to react with the intermediate phosphonium compound, in a manner quite analogous to that of the halide ion in the classical Michaelis-Arbuzov reaction (eq. i), to yield the phosphorothioate and a thioether. Symmetrical dialkyl disulfides react at about 140° and diaryl disulfides at 80°, but with mixed alkyl aryl disulfides the reaction takes place exothermi-cally at room temperature. The reaction of sodium diethyl phosphonate, even with symmetrical dialkyl disulfides, takes place instantaneously at o° to yield the phosphorothioate and sodium alkylmercaptide. If these two products are not separated immediately, they interact further, as indicated in equation n, to form a thioether and the salt of the phosphorothioate.

It is noteworthy that in the reactions of both equation 10 and equation 11, the attack of the mercaptide anion is on a C-O-P rather than a C-S-P linkage. The marked reactivity of mercaptide anion in this cleavage reaction, together with the observation that S-p-chlorobenzylthiouronium salts afford crystalline derivatives of phosphonic, phosphoric, and phos-phorothioic acids, suggested a convenient procedure for the positive identification of phosphate, phosphonate, and phosphorothioate esters, substances which usually are liquids and often difficult to obtain in analytically pure form. Treatment of the ester with sodium ethyl mercaptide smoothly cleaves one alkoxy linkage to yield a monobasic sodium salt which is readily converted to the sharp melting S-p-benzylthiouronium derivative (26, d7).