PT-141

The seven residues, numbered from the N-terminus, are Nle1, Asp2, His3, D-Phe4, Arg5, Trp6 and Lys7. An amide bond between the side-chain carboxyl of Asp2 and the side-chain amine of Lys7 closes a six-residue ring, Asp2 to Lys7, and the N-acetyl-norleucine cap sits outside it. Norleucine, 2-aminohexanoic acid, is an unbranched isomer of leucine and not a coded amino acid.

Forming the lactam eliminates one molecule of water, so the ring is 18 Da lighter than the open chain. D-Phe4 is the one D-configured residue; a diastereomer with L-Phe in its place has exactly the same mass, which is why HPLC retention, not mass alone, tells the two apart.

His3, D-Phe4, Arg5 and Trp6 sit inside the ring with Asp2 and Lys7, so the four side chains that differ most in shape, imidazole, phenyl, guanidine and indole, are all held within the lactam.

PT-141 ends in a free carboxylic acid on Lys7, written -OH, while the N-terminus is acetylated. The formula C50H68N14O10 has one oxygen more and one nitrogen and one hydrogen fewer than an amide-terminated chain of the same residues, a monoisotopic difference of 0.98 Da.

That free carboxyl carries a negative charge near pH 7, which offsets most of the positive charge of Arg5. The calculated net charge near pH 7 is about +0.2, and the C-terminal acid also shifts the peptide's retention in reversed-phase HPLC relative to the amide form.

With the N-acetyl cap leaving no free amine at the start of the chain and Lys7 used in the lactam, Arg5 and the partly protonated His3 are the only positive groups.

The monoisotopic mass is 1024.5243 Da, so the [M+H]+ ion is expected at m/z 1025.5316 and the [M+2H]2+ ion at m/z 513.2694. Resolving that value from the amide form, about 1 Da lower, needs a mass spectrometer that separates isotope peaks cleanly, and the HPLC retention time adds a second, independent check.

Tandem mass spectrometry needs two backbone cleavages to free a fragment from inside the ring, so the cyclic chain gives a fragment pattern unlike a linear chain of the same residues, which confirms that the lactam is closed.

Trp6 absorbs at 280 nm, and the peptide bonds around 214 nm, so reversed-phase HPLC gives purity as the main peak's share of total peak area at either wavelength. The material is a white to off-white lyophilised solid in a sealed vial. The sealed vial is stored at 2–8 °C short term, or at −20 °C and below for long-term storage, protected from light and away from repeated freeze-thaw cycles.