Thiohydantoins, formation

Thioamide formation benzodiazepinone, 505 heteiodiazepinone, 621 phosphorus pentasulf ide, 323, 600 Thioazole formation, nitrile addition, 301 Thiocarbamate formation, 588 phenol, 95 rearrangement, 517 Thioenol ether formation, 185, 517 addition-elimination, 554 Thioester formation, mixed anhydride, 184 Thioether formation, 241, 300, 413, 416 alkylation, 586, 588 aromatic displacement, 416 Thiohydantoin formation, 293 Thiol interchange, benzothiazole formation, 422... 

To increase the number of diversities, the hydantoin (or thiohydantoin) formation reaction was performed starting from N-alkylated dipeptides (Fig. 3). In the last synthesis step, the hydantoin (or thiohydantoin) ring was alkylated followed by the cleavage from the resin. Using 54 different amino acids for the first position of diversity (R1), 60 different amino acids for the second position of diversity (R2), and four different alkylating... 

In the experiments of Haurowitz et al. (1957) proteins and peptides were in an acetic anhydride/acetic acid medium during thiohydantoin formation. Their results with a-peptides and poly-a-glutamic acid indicated that very little a —> 7 interconversion took place, but they accepted that it could occur if glutamic acid residues were adjacent to certain other amino acid residues. These authors also suggested that possible y a conversion... 

Although further modification, e.g., substituted thiohydantoin formation, may result under other circumstances, thiourea derivatives are ordinarily quite stable near neutrality. 

The formation of a sulfur-containing ring was justified by the attack of the halogenated carbon of the cMoroacetyl derivative by the sulfur atom of thiourea, a fact in accordance with the results just discussed by Wallach (49,50) and Claus (51). The new formula (37) of thiohydantoine explained why, contrary to thiourea, its desulfuration was difficult. 

Mass spectral fragmentation patterns of alkyl and phenyl hydantoins have been investigated by means of labeling techniques (28—30), and similar studies have also been carried out for thiohydantoins (31,32). In all cases, breakdown of the hydantoin ring occurs by a-ftssion at C-4 with concomitant loss of carbon monoxide and an isocyanate molecule. In the case of aryl derivatives, the ease of formation of Ar—NCO is related to the electronic properties of the aryl ring substituents (33). Mass spectrometry has been used for identification of the phenylthiohydantoin derivatives formed from amino acids during peptide sequence determination by the Edman method (34). 

The mechanism involves the initial formation of a substituted urea followed by ring closure to form the thiohydantoin. The amino acid is dissolved in 60% aqueous pyridine containing the phenylisothiocyanate... 

A-Thiazolyl a-amino acids 56 have been prepared. The preferred route to these compounds would utilise the Hantzsch synthesis, however in this case the in situ formation of the required thiourea derivatives of a-aminoacids 52 failed. A variety of isothiocyanate reagents were tried, with the result being either no reaction, decomposition or the corresponding thiohydantoin 53. A modified version of the Hantzsch synthesis was developed. If the bromoketone 54 is initially treated with sodium thiocyanate an a-thiocyanatoketone 55 is formed, subsequent addition of the amino acid ester 51 yields A-thiazolyl a-amino acids 56 <00T3161>. 

Fairwell, T., S. Ellis, and R.E. Lovins, Quantitative protein sequencing using mass spectrometry thermally induced formation of thiohydantoin amino acid derivatives from N-methyl- and N-phenylthiourea amino acids and peptides in the mass spectrometer. Anal Biochem, 1973. 53(1) 115-23. 

Microwave-induced imine formation, subsequent reduction with NaBH(OAc)3 and cyclisation of the resulting amino acid with isothiocyanates was used in an efficient one-pot multi-step synthesis of thiohydantoins (Scheme 4.26). The reductive animation was conducted as a two-step procedure to avoid direct reduction of the aldehyde at high temperatures48. 

PITC). The mixture is warmed at 40 °C for 1 h, then diluted with 1 ml of water and the excess of MITC (or PITC) is removed by extracting four times with 2-ml volumes of benzene. The aqueous layer is evaporated and dried in a vacuum desiccator over sodium hydroxide. The general scheme for the formation of PTH-amino acids is illustrated in Fig.4.1. For peptide hydrolysis, 1.5 ml of a mixture consisting of equal volumes of 3 N hydrochloric acid and 60% acetic acid are added and the reaction is incubated in a nitrogen atmosphere for 30 min at 40 °C. The mixture is diluted with 2 ml of water and the thiohydantoin derivative is extracted with 2 ml of ethyl acetate followed by 2 ml of benzene. The combined extracts are used for chromatography. 

Thiohydantoins (see Scheme 4.25, p. 78) play an important role as derivatives of amino acids, particularly in the sequential analysis of peptides. When the sequence is being determined from the carboxyl end, ammonium thiocyanate is dissolved in acetic acid and acetic anhydride and this mixture is allowed to react with the carboxyl end of the peptide with the formation of l-acyl-2-thiohydantoin. 2-Thiohydantoin is released from the peptide by the action of a base, and a new carboxyl end of the amino acid is exposed. Prior to GC analysis, thiohydantoins must be further modified, e.g., by silylation [271], as follows. A 25-/A volume of ethyl acetate and 25 /d of BSA are added to 500 nmol of 2-thiohydantoin and the mixture is shaken until it is homogeneous. The mixture is then heated in a stoppered test-tube at 80°C for 5 min, cooled and centrifuged before injection. 

The inability of C-terminal proline to be derivatized to a thiohydantoin has been a major impediment to the development of a routine method for the C-terminal sequence analysis of proteins and peptides. Since the method was first described in 1926 (2), the derivatization of C-terminal proline has been problematic. While over the years a few investigators have reported the derivatization of proline, either with the free amino acid or on a peptide, to a thiohydantoin (6-8), others have been unable to obtain any experimental evidence for the formation of a thiohydantoin derivative of proline (9-12). Recently, utilizing a procedure similar to that described by Kubo et al. (6), Inglis et al. (13) have described the successful synthesis of thiohydantoin proline from N-acetylproline. This was done by the one-step reaction of acetic anhydride, acetic acid, trifluoroacetic acid, and ammonium thiocyanate with N-acetyl proline. We have reproduced this synthesis and further developed it to a large scale synthesis of TH-Proline. 

Use, D., and Edman, P., The formation of 3-phenyl-2-thiohydantoins from phenylthiooarbamoyl amino acids. Aust. J. Chem. 16, 411-416 (1963). 

Chang, J.-Y, A novel Edman-type degradation direct formation of the thiohydantoin ring in alkaline solution by reaction of Edman-type reagents with N-monomethyl amino acids, FEBS Lett., 91, 63-68, 1978. 

In contrast, reaction of 5,5-diphenyl thiohydantoin with l-bromo-2-chloroethane generates two isomeric imidazo(2,l-b)thiazole derivatives (97 and 98) through intramolecular S,N-dialkylation.235 The formation of imidazo(2,l-b)thiazoles by this kind of process is rather common.236 Similarly, reaction with 1,3-dibromopropane gives two isomeric diphenylimidazothiazines.2 3 7,2 3 8... 

Hydantoins and thiohydantoins have been used as analytical reagents for heavy metals because of their capacity for complex formation.361-363... 

The kinetics and mechanism of the formation of 3,5-disubstituted 2-thiohydantoins were described by Drobnica and Augustin [77, 78], It is thought that during the cooking or processing of food and also in the human digestive system, isothiocyanates in foods react partly with free amino acids to form their corresponding 2-thiohydantoins. 

Fig. (lO). Formation of 3,5-disubstituted 2-thiohydantoins from MTBI and amino acids. 

Trimethylsilylation of a number of thiohydantoins gave products (silylated at both nitrogen atoms) with generally good GC characteristics a complication was the formation of tris-silylated byproducts, created by additional silylation of the enol form of the thiohydantoin. 

Similarly, thiohydantoins 29 were prepared by using thiophosgene or thiocarbonyldiimidazole as the reagent. A combinatorial library containing 6364 individual thiohydantoins was then assembled in the positional scanning format. For this library, 74 and 86 amino acids were used in the Rj and R3 positions, respectively. 

A series of l-oxo-3-thioxo-2,3,7,7a-tetrahydro-l/f-pyrrolo[l,2-c]imidazoles (133) have been obtained by reaction of 2-thiohydantoin with activated alkenes. The formation of this pyr-roloimidazole can be explained by an initial addition of 2-thiohydantoin to the activated double bond to yield an acyclic intermediate with subsequent cyclization (Equation (10)) <828502,83LA1468). 

Imidazoline-2-thione derivatives such as 43 were synthesized by condensation of 1-amino-l-deoxy-D-fructose (or its iV-alkylated derivatives) with glycosyl isothiocyanates and converted to dehydration products or spiro-cyclic derivatives such as 44 and 45, respectively, under different acidic conditions. The spiro-thiohydantoin analogues 47 and 48 were synthesized from the photo-bromination product 46 (Scheme 9), the latter being able to interfere with the formation of glucose from glycogen in rats. ... 

Only the mechanism for the formation of peptidyl 2-thiohydantoin is illustrated here. 

This reaction has been extended to the reaction of terminal amino acids with thiocyanic acid7b,9 developed initially by Johnson ) and trimethylsilyl isothiocyanate for the formation of thiohydantoin. In addition, 12 N HCl and acetohydroxamate have been used to cleave the 2-thiohydantoin from the peptide chain. 

Fluorescein isothiocyanate was proposed by Maeda el al. as a fluorescent end-group reagent [218] analogous to phenylisothiocyanate in the Edman method. A procedure for the determination of free amino acids by formation of fluorescent thiohydantoins with fluorescein isothio-cytinate was reported by Kawauchi ef al. [219]. 

Figure 9. Formation of fluorescent thiohydantoins by reaction of an isothiocyanate with an amino acid. X = fluorophore.

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