[87-86-5]  · C6HCl5O  · Pentachlorophenol  · (MW 266.32)

(coupling reagent in peptide synthesis, to form active esters; a probe in various NQR studies on pentachlorophenol-amine hydrogen bonded complexes;1 a standard in GC analysis2

Physical Data: mp 188-191 °C (anhyd); bp 310 °C; d 1.978 g cm-3.

Solubility: sol ether, alcohol, hot benzene, ligroin; insol H2O.

Form Supplied in: widely commercially available.

Purification: recrystallize from alcohol (monoclinic prisms) or benzene (needles) and vacuum dry.

Handling, Storage, and Precautions: highly toxic and possible teratogen. May be fatal if inhaled, swallowed, or absorbed through the skin. Causes skin and eye irritation and irritation to the mucous lining of respiratory tract. Do not breathe dust; avoid prolonged exposure. Readily absorbed through the skin. Incompatible with strong oxidizing agents, strong bases, acid chlorides, and anhydrides. More basic in organic solvents. Store in a cool dry place. There are concerns as to its environmental chemical effects.2,3

Polypeptide Synthesis.

The most important stage in peptide synthesis is the coupling step of one amino acid or peptide to another. Coupling reagents have been proposed following early attempts at coupling because of their inherent convenience. These generally work by peptide bond formation between a carboxyl and an amine component in the presence of a specific condensing or coupling agent. In the late 1950s and early 1960s, considerable effort was made to synthesize polypeptides with a known repeating sequence of amino acids. The problem until then with the more traditional methods4a,b (e.g. mixed anhydrides and the 1,3-Dicyclohexylcarbodiimide method) was undesirable side reactions4 which could lead to early termination of polymerization; in addition, various other methods which avoid this, e.g. thiophenyl and p-nitrophenyl esters, may not be used when N-protecting groups are removed by catalytic hydrogenation.4,5 The use of pentachlorophenyl esters was first fully reported in 19656 although there had been brief references to their use before then.7a,b

The use of pentachlorophenyl active esters in the synthesis of polypeptides of known repeating amino acid sequence has several advantages:

  • 1)Pentachlorophenyl esters work particularly efficiently when coupled with benzyloxycarbonyl and t-butyl protecting groups in trifunctional amino acids for polymerization, e.g. a protecting N-Cbz group is selectively cleaved by catalytic hydrogenation,6 leaving pentachlorophenyl and t-butyl groups intact. Compare HBr cleavage of the Cbz group which would prevent the use of t-butyl groups (t-butyl groups are essential when utilizing aspartic or glutamic acids to prevent trans-peptidation reactions).
  • 2)Pentachlorophenyl esters of both amino acids and peptides tend to be easier to purify since they generally have higher melting points compared to other active esters. This assists in the purification of C- or N-activated peptide intermediates which must be very pure in polymerization to avoid early termination reactions. Table 18 gives some illustrative examples of this, along with melting point values of other active esters for comparison.

    Given these requirements, pentachlorophenyl esters give polypeptides of higher molecular weights than those obtained by other methods, as determined by end-group analysis and viscometry.

    Pentachlorophenyl active esters are amongst the most reactive esters, e.g. aminolysis of Cbz-L-phenylalanine pentachlorophenyl ester has a half life of 1.34 min on treatment with benzylamine in dioxane; cf. the half life of the 2,4,5-trichlorophenyl ester which is 4.9 min and the p-nitrophenyl ester which is 23.2 min.8 This short reaction time limits the time available for side reactions to occur during coupling reactions.

    The first cited example of pentachlorophenyl esters being utilized was in the synthesis of poly-b-L-aspartic acid.9 This polymer was required in optically pure form for various chemical, biological, and immunochemical investigations.10 In summary, the coupling of the amino acid with pentachlorophenol using the DCC method (see below) gives an active ester which polymerizes rapidly in the presence of a tertiary amine to give a poly(amino acid) of higher molecular weight than is obtained using other methods, notably the p-nitrophenyl ester.

    Another example is the synthesis of poly-b-L- (or -D-) glutamyl-b-alanine11 (3) from the precursor a-t-butylglutamyl-b-alanine pentachlorophenyl ester hydrochloride (2).

    The precursor (2) was prepared according to eq 1. N-Benzyloxycarbonyl-a-t-butylglutamic acid pentachlorophenyl ester was coupled to the methyl ester of b-alanine to give (4a). This was hydrolyzed using NaOH to give the free acid (4b) which was subsequently coupled to pentachlorophenol with DCC to give N-benzyloxycarbonyl-a-t-butylglutamyl-b-alanine pentachlorophenyl ester (4c; 60% yield, >97% optically pure). The same method was used to prepare the 2,4,5-trichloro- (4d) and p-nitrophenyl (4e) esters for comparison, but the pentachlorophenyl esters immediately proved easier to purify, for the reasons already described.

    Hydrogenation of (4c) using 10% Palladium on Carbon catalyst in the presence of HCl in methanol gave (2) in good yield (59%). Polymerization in DMF with an excess of Triethylamine at the highest possible concentrations gave (3) with t-Bu protecting groups which were readily removed with 90% Trifluoroacetic Acid to give the free polypeptide.

    There are many other examples of pentachlorophenyl active esters being used in peptide syntheses similar to those described above, but these will not be described in detail here.8,12a-f

    Very little by way of modification or variation in pentachlorophenyl active ester coupling procedures has occurred; they have mostly found new applications. There is, however, one exception where a different procedure has been used to prepare pentachlorophenyl esters of N-protected amino acids.13 The pentachlorophenyl di- (5) or tri- (6) chloroacetates (known plant growth regulators) were prepared and subjected to ester exchange reactions with trialkylammonium salts of acylamino acids (7) in DMF, rapidly giving good yields of the desired esters (eq 2).

    One last novel reaction which may involve pentachlorophenyl esters is the formation of polypeptides and Leuchs' anhydrides from carbamic acid derivatives of C-activated amino acids.14 C-Activated N-Cbz amino acids (8) on catalytic hydrogenation give the carbamic acid derivative (9), which cyclizes to the corresponding Leuchs' anhydride (10) (eq 3); in some cases this immediately polymerizes to the polypeptide, e.g. the mixed anhydride N-Cbz-DL-phenylalanine polymerized to poly-L-phenylalanine via the Leuchs' anhydride.

    1. (a) Grech, E.; Kalenik, J.; Sobczyk, L. JCS(F1) 1979, 75, 1587. (b) Kalenik, J.; Majerz, I.; Malarski, Z.; Sobczyk, L. Chem. Phys. Lett. 1990, 165, 15.
    2. van Langeveld, H. E. A. M. J. Assoc. Off. Anal. Chem. 1975, 58, 19.
    3. Insecticide for termite control; pre-harvest defoliant and general herbicide, it has been recommended for use in the preservation of wood and wood products, starches, dextrins and glues. There are many examples of the environmental and toxic effects of pentachlorophenol, e.g. (a) Uhl, S.; Schlatter, C.; Schmid, P. Arch. Toxicol. 1986, 58, 182. (b) Uhl, S.; Schlatter, C.; Schmid, P. Rev. Environ. Contam. Toxicol. 1988, 104, 184. (c) Seiler, J. P. Mutation Res. 1991, 257, 27.
    4. (a) Goodman, M.; Kenner, G. W. Adv. Protein Chem. 1957, 12, 465. (b) Albertson, N. F. OR 1962, 12, 157.
    5. Kovacs, J.; Ballina, R.; Rodin, R. CI(L) 1963, 1955.
    6. Kovacs, J.; Kapoor, A. JACS 1965, 87, 118.
    7. Few earlier references to pentachlorophenyl esters except for (a) Kupryszevski, G. Rocz. Chem. 1961, 35, 1533 (preparation of a few derivatives). (b) Stich, K.; Lehmann, H. G. HCA 1963, 46, 1887. (c) Pless, J.; Boissonnas, R. H. HCA 1963, 64, 1609 (studied the kinetics of aminolysis of Cbz-Phe pentachlorophenyl active ester).
    8. Kovacs, J.; Giannotti, R.; Kapoor, A. JACS 1966, 88, 2282 and references therein.
    9. Kovacs, J.; Ballina, R.; Rodin, R. L.; Balasubramanian, D.; Applequist, J. JACS 1965, 87, 119.
    10. Kovacs, J.; Ballina, R.; Rodin, R. CI(L) 1963, 1955.
    11. Kovacs, J.; Johnson, B. J. JCS 1965, 6777.
    12. (a) Kovacs, J.; Kisfaludy, L.; Ceprini, M. Q. JACS 1967, 89, 183. (b) Katakai, R.; Goodman, M. Macromolecules 1982, 15, 25. (c) Bodanszky, M. Int. J. Pept. Protein Res. 1985, 25, 449. (d) Bodanszky, M.; Bodanszky, A. The Practice of Peptide Synthesis; Springer: Berlin, 1984; p. 120. (e) Johnson, B. J. JPS 1973, 62, 1019. (f) Chemical Abstracts reveals many more examples, which are far too numerous to include here.
    13. Fujino, M.; Hatanaka, C. CPB 1968, 16, 929.
    14. Kovas, J.; Kovacs, H. N.; Chakrabarti, J. K.; Kapoor, A. E 1965, 21, 20.

    Adrian P. Dobbs

    King's College London, UK

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