Amino isothiocyanate coupling

Special chemical derivatives free thiol functions of cysteine can be problematic because of post-synthetic uncontrolled oxidation. To avoid this, you can replace Cys by serine (Ser), alanine (Ala), or u-aminobutyric acid (Abu). Alternatively, choose the hydrophilic Cys(Acm) and leave protected. For the simultaneous preparation of peptides of different size with free amino terminus, couple the terminal amino acid as /V-Boc derivatives so that they will not become acetylated during the normal elongation cycle. Boc is removed during the final side chain deprotection procedure (Protocol 4). Special labels can be attached to the N-termini by spotting respective derivatives in an additional coupling cycle. We have succes lly added biotin via the in situ formed HOBt-ester (normal activation procedure) or fluorescein via the isothiocyanate (FITC) dissolved in NMP. 

Obviously, the main purpose for the introduction of CL detection coupled to CE separations is inherent to the development and improvement of sensitive and uncomplicated devices to achieve a decrease of the band broadening caused by turbulence at the column end, together with the attractive separation efficiency of CE setups. With this purpose in mind, Zhao et al. [83] designed a postcolumn reactor for CL detection in the capillary electrophoretic separation of isoluminol thiocarbamyl derivatives of amino acids, because, like other isothiocyanates, isoluminol isothiocyanate has potential applications in the protein-sequencing area. 

Ricci and co-workers introduced a new class of amino- alcohol- based thiourea derivatives, which were easily accessible in a one-step coupling reaction in nearly quanitative yield from the commercially available chiral amino alcohols and 3,5-bis(trifluoromethyl)phenyl isothiocyanate or isocyanate, respectively (Figure 6.45) [307]. The screening of (thio)urea derivatives 137-140 in the enantioselective Friedel-Crafts reaction of indole with trans-P-nitrostyrene at 20 °C in toluene demonstrated (lR,2S)-cis-l-amino-2-indanol-derived thiourea 139 to be the most active catalyst regarding conversion (95% conv./60h) as well as stereoinduction (35% ee), while the canditates 137, 138, and the urea derivative 140 displayed a lower accelerating effect and poorer asymmetric induction (Figure 6.45). The uncatalyzed reference reaction performed under otherwise identical conditions showed 17% conversion in 65 h reaction time. 

The N-terminal amino group or a Lys residue can easily be used to label the PNA. This can be carried out in solution after purification, or more conveniently, while the PNA is still attached to the resin. Fluorescein and rhodamine are the most common fluorophore labels, while biotin is the most common affinity tag. Fluorescein or rhodamine are usually coupled to the amino group as -OSu esters or isothiocyanates. 4,4 -Dimethoxytrityl-protected biotin and Piv-pro-tected fluorescein have also been coupled to the N-terminus of a PNA as their 1-phenylpyr-azolin-5-one carboxylate esters.147 In 1997 a new fluorescein-conjugated Lys monomer (21, Scheme 12) was described. 48 ... 

This reaction takes place because diimides, —N=C=N—, have reactive cumulated double-bond systems like those of ketenes, C=C=0 isocyanates, —N=C=0 and isothiocyanates, —N=C=S and are susceptible to nucleophilic attack at the central carbon. In the first step of the diimide-coupling reaction, the carboxyl function adds to the imide to give an acyl intermediate, 9. This intermediate is an activated carboxyl derivative RCO—X and is much more reactive toward an amino function than is the parent acid. The second step therefore is the aminolysis of 9 to give the coupled product and yV,N -dicyclohexylurea ... 

The isothiocyanate derivative of the fluorophore, fluorescein or rhodamine, is coupled to the amino groups of IgG antibody in a one-step procedure and excess label is removed by gel filtration. 

Fig. 20. Direct coupling of glyconolactones (31) and sugars activated as isothiocyanates (29) to dendrimers bearing primary amino groups.

Coupling of the amino alcohol d-48 and the isothiocyanate 7 in 75% aq. DMF gave the thiourea derivative d-401 (100%), which was treated with an excess of yellow mercury(II) oxide in diethyl ether to afford d-402 (93%). Removal of the benzyl ether groups by sodium in liquid ammonia was followed by purification by a column of Dowex 50W-X2 (H+) resin to give the trehazolin analogue d-403 (90%). The hepta-N,O-acetyl derivative d-404 fully supported the structures assigned for d-403 (Scheme 52).118... 

The amino alcohols d-50 and l-50 were converted into d-408 and l-408 via coupling with isothiocyanate 7 to give the thiourea d-405 (93%), which was similarly converted into the cyclic isourea d-406 100%) (Scheme 53).98 Conventional O-debenzylation afforded the trehazolin analogue d-407 (96%), which was further characterized as the octa-iV,O-acetyl derivative d-408. Likewise, the diastereoisomer l-407 was synthesized in 91% overall yield by similar reaction sequence, starting from the l-405 (97%) obtained from isothiocyanate 7 and aminotetrol l-50. 

Hydrolysis of the oxazolidinone 236 gave the corresponding amino alcohol, which without isolation was coupled with the sugar 4-isothiocyanate 2653 to provide the thiourea 452 (Scheme 67).105 Cyclization of 452 to the isourea 453 was carried out with yellow mercuric oxide, and the resulting oxazoline 453 was debenzylated to give... 

Amino-5-methyltriazole and carbethoxy isothiocyanate, stirred in acetonitrile, gave 3-methyl-5-thioxo-l,2,3-triazolo[l,5-a][l,3,5]triazin-7-pne (104) (25°C, 30 min, 65%) (76JHC589). 5-Phenyltriazole-4-diazonium chloride and phenyl isocyanate, stirred in dichloromethane, provided 3,6-diphenyl-l,2,3-triazolo[5,l-d][l,2,3,5]tetrazine-4-one (105) (25°C, 41%) (79TL4253). Finally, diazotized 4-aminotriazole was coupled to 2-naphthol. The product, refluxed in methanol, was cyclized to naphtho[2,l-e][l,2,3]-triazolo[l,5-h] triazine (106) (2 days, 80%) (74JHC867). 

Most amino acids react with ninhydrin at ambient temperatures to form a blue color that becomes purple on heating. However, proline and hydroxyproline yield yellow compounds that are measured at a different wavelength. Other postcolumn derivatizations use fluorogenic reagents, such as o-phthaldialdehyde or fluorescamine. Precolumn derivatization techniques using o-phthaldialdehyde, dansyl, phenyl isothiocyanate, or 9-fluorenylmethyl chloroformate derivatives have been used with reversed-phase HPLC. Electrochemical detection has also been coupled with derivatization methods to enhance analytical sensitivity. 

The amino group of 4-amino-3-pyrazolin-5-ones can be diazotized easily and the resulting diazonium salt couples with the usual reagents. Other diazonium salts couple with 4-amino-3-pyrazolin-5-ones at the amino group. Mercuric halides replace the amino hydrogen in 2,3-dimethyl-l-phenyl-4-sulfamino-3-pyrazolin-5-one.543 The 4-amino groups react normally with isothiocyanates,49 epoxides,1261 ureas1367 and nitrosamines.527... 

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