Determination of non-protein α-alkyl-α-amino acids by reversed-phase high-performance liquid chromatography in comparison with ion-exchange and capillary gas chromatography

Brückner, Hans; Bosch, I.; Graser, Theodor; Fürst, Peter

doi:10.1016/s0021-9673(01)94602-3

Cited by 17 publications

(3 citation statements)

References 49 publications

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“…However, Aib and C°-MeLeu react considerably more slowly than the -amino acids and would need a 10-and 15-min reaction time, respectively, to reach their maximum fluorescence. 41 As a result, Aib gave a very low but quantifiable signal, while C"-MeLeu could not be detected in our system. A precolumn derivatization would appear to be necessary, but we envisage that other inefficiencies would result due to the relative instability of some OPA-adducts like OPAglycine.41 So, amino acid analysis of C°-methyl amino acid-containing peptides was coupled to mass spectrometry to confirm their structure (see Table I).…”

Section: Resultsmentioning

confidence: 68%

“…This results in very low color yields with ninhydrin (under standard conditions, ninhydrin color yield of Aib and C"-MeLeu relative to Leu are 7.4 and 9.4%, respectively, as determined by Brückner et al). 41 This property results in less accurate monitoring of the subsequent coupling step by the Kaiser test. 42 We systematically repeated this particular step three times with large excesses of Boc-amino acid and DIC and then deblocked an aliquot of the resin with TFA.…”

Section: Resultsmentioning

confidence: 99%

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Synthesis and relative potencies of new constrained CRF antagonists

Hernandez¹,

Kornreich²,

Rivier³

et al. 1993

J. Med. Chem.

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Two series of CRF antagonists with N alpha- and C alpha-methylated alanine and leucines were evaluated for their biological activities in vitro and in vivo in several systems. The poly-N-methylated analogue of alpha-helical-CRF9-41, [N alpha MeLeu10,15,27,37,N alpha MeAla22,32,41]-alpha-Hel-CRF9-41, was found to be considerably less potent than the parent non-N-methylated analogue. This result was expected on the basis that alpha-helicity was thought to be required for biological activity and the prediction that backbone substitutions on the nitrogen have a tendency to break alpha-helices (a hypothesis that was confirmed by circular dichroism). Next, a series of constrained analogues of the potent CRF antagonist, [DPhe12,Nle21,38]h/rCRF12-41, was synthesized that contained C alpha-methylleucine and/or C alpha-methylalanine (Aib) residues at selected positions. Because C alpha-methylation is recognized to increase alpha-helicity, and because there is now strong NMR data suggesting that residues 6-36 assume a well-defined alpha-helix, it was expected that these analogues would be more potent. Although usual solid-phase peptide synthesis procedures were followed, success in coupling the C alpha-methyl amino acids was obtained only with a 1:1 mixture of BOP/HOBt. In vitro potencies of the synthesized compounds were measured in a collagenase-dispersed anterior pituitary cell culture bioassay. Monosubstituted analogues were shown to be twice to one fourth as potent as the parent compound; while the pluri-substituted peptides were slightly less potent. This decrease in potency might be correlated to an unexpected lower helical content of the pluri-substituted compounds (as determined by CD spectroscopy), as it was suggested that the bioactive conformation of the CRF was predominantly alpha-helical. Interestingly, one analogue, [DPhe12,Nle21,38,C alpha-MeLeu37]h/rCRF12-41, was found to be more potent and longer acting than the parent compound in two in vivo assays measuring ACTH release after intravenous administration to adrenalectomized rats and reversal of stress-induced delay in gastric emptying in the rat after intracisternal administration. The molecular basis for this increased duration of action and potency is being investigated.

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Section: Resultsmentioning

confidence: 68%

Section: Resultsmentioning

confidence: 99%

Synthesis and relative potencies of new constrained CRF antagonists

Hernandez¹,

Kornreich²,

Rivier³

et al. 1993

J. Med. Chem.

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“…Highly sensitive determination of <-(7-glutamyl)lysine cross-links has been performed by liberation of the crosslink by exhaustive proteolytic digestion and derivatization with o-phthalaldehyde (OPA) followed by highperformance liquid chromatography (HPLC) separation (Griffin et al, 1982;Beninati et al, 1988; Nonaka et al, 1989; Tanaka et al, 1990;Kimura et al, 1991). It has been, however, demonstrated that the fluorescence intensity of OPA-2-mercaptoethanol derivatives depends on temperature and reaction time (Brückner et al, 1987). Moreover, the relative fluorescence of 0PA-e-(7-glutamyl)lysine decreases to 83 % of its initial value after only 45 min in storage (Griffin et al, 1982).…”

Section: Introductionmentioning

confidence: 99%

Improved method for identification and determination of .epsilon.-(.gamma.-glutamyl)lysine cross-link in protein using proteolytic digestion and derivatization with phenyl isothiocyanate followed by high-performance liquid chromatography separation

Satô¹,

Tsukamasa²,

Imai³

et al. 1992

J. Agric. Food Chem.

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Sequences of Metanicins, 20‐Residue Peptaibols from the Ascomycetous Fungus CBS 597.80

Kimonyo

Brückner

2013

Chemistry & Biodiversity

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Four linear 20-residue peptaibols, named metanicins (MTCs) A-D, were isolated from submerged cultures of the ascomycetous fungus CBS 597.80. Structure elucidation was performed by a combination of fast-atom-bombardment mass spectrometry (FAB-MS), electrospray ionization MS, Edman degradation of isolated fragments, and amino acid analysis by ion-exchange and gas chromatography, and enantioselective HPLC. The sequences of MTC A(B) are (amino acid exchange in B and C in parentheses): Ac-Aib-Ala-Aib-Ala-Aib-Ala-Gln-Aib-Val-Aib-Gly-Leu-Aib-Pro-Val-Aib-Aib(D-Iva)-Gln-Gln-Pheol and of MTC C(D) Ac-Aib-Ala-Aib-Ala-Aib-Ala-Gln-Aib-Val-Aib-Gly-Leu-Aib-Pro-Val-Aib-Aib(D-Iva)-Gln-Gln-Pheol (Ac, acetyl; Aib, α-aminoisobutyric acid; Iva, isovaline; Pheol, L-phenylalaninol). The peptides are related, and some of the sequences are identical, to other 20-residue peptaibols isolated from Trichoderma species. MTCs show moderate activities against Micrococcus luteus, Enterococcus faecalis, and Staphylococcus aureus, and very low activities against Bacillus subtilis. The producer has originally been identified and deposited as Metarhizium anisopliae var. anisopliae CBS 597.80. Although this identification has been withdrawn by Centralbureau voor Schimmelcultures (CBS) in the meantime, the accession number will be retained - independently from any taxonomic revisions.

show abstract

Determination of non-protein α-alkyl-α-amino acids by reversed-phase high-performance liquid chromatography in comparison with ion-exchange and capillary gas chromatography

Cited by 17 publications

References 49 publications

Synthesis and relative potencies of new constrained CRF antagonists

Synthesis and relative potencies of new constrained CRF antagonists

Improved method for identification and determination of .epsilon.-(.gamma.-glutamyl)lysine cross-link in protein using proteolytic digestion and derivatization with phenyl isothiocyanate followed by high-performance liquid chromatography separation

Sequences of Metanicins, 20‐Residue Peptaibols from the Ascomycetous Fungus CBS 597.80

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