The Enantioselective Immunoaffinity Extraction of an Optically Active Ibuprofen-Modified Peptide Fragment

Ikegawa, Shigeo; Isriyanthi, Ni Made Ria; Nagata, Masanori; Yahata, Kenji; Ito, Hiromi; Mano, Nariyasu; Goto, Junichi

doi:10.1006/abio.2001.5198

Cited by 14 publications

(10 citation statements)

References 32 publications

(33 reference statements)

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“…By employing phosphate buffered saline (PBS), pH 7.4, as the sole mobile phase, we were able to reuse the same antibody-columns for more than 2000 separations over a period of 3 years. This is in contrast to various other reports that demonstrated separation of stereoisomers in immunoaffinity systems [4][5][6][7][8][9]. In those studies, mixtures of enantiomers, dissolved in a neutral buffer, were passed through a column containing antibodies covalently linked to agarose beads [4][5][6][7][8][9] or silica [9]; after the unbound enantiomer was washed clear, elution of the bound enantiomer was achieved by altering to a mobile phase that disrupted its interaction with the antibody.…”

Section: Introductionmentioning

confidence: 92%

“…In those studies, mixtures of enantiomers, dissolved in a neutral buffer, were passed through a column containing antibodies covalently linked to agarose beads [4][5][6][7][8][9] or silica [9]; after the unbound enantiomer was washed clear, elution of the bound enantiomer was achieved by altering to a mobile phase that disrupted its interaction with the antibody. As is typical for classical affinity and immunoaffinity chromatography systems [10][11][12], elution of bound material required harsh conditions, i.e., a drastic change in pH [9], or addition of organic solvents [4,[6][7][8] or chaotropic salts [5]. Such severe elution conditions invariably cause protein denaturation and considerably shorten column lifetime, which rarely exceeded 100 separation cycles [9].…”

Section: Introductionmentioning

confidence: 99%

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Effect of the mobile phase on antibody-based enantiomer separations of amino acids in high-performance liquid chromatography

Hofstetter

Lindstrom

Hofstetter

2004

Journal of Chromatography A

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The effect of the mobile phase parameters flow rate, temperature, pH and ionic strength, as well as the addition of various organic modifiers on the enantiomer separation of various aromatic alpha-amino acids was investigated using two antibody-based chiral stationary phases that have opposing stereoselectivity. On both columns, a decrease in flow rate or temperature resulted in increased interaction with the retained enantiomer. It was found that the retention factor k2 depends on the affinity between the analyte and the immobilized antibody and is not independent of the flow rate. Optimum separations of all amino acids investigated were obtained at pH 7.4 on both columns. While increased k2 values were obtained at low ionic strength on the anti-D-amino acid antibody column, no such effect was observed on the anti-L-amino acid antibody column. The addition of organic modifiers did not improve separations. In all studies, the unretained enantiomer eluted with the void volume.

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Section: Introductionmentioning

confidence: 92%

Section: Introductionmentioning

confidence: 99%

Effect of the mobile phase on antibody-based enantiomer separations of amino acids in high-performance liquid chromatography

Hofstetter

Lindstrom

Hofstetter

2004

Journal of Chromatography A

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“…One potential application for these IA-RAM columns would be their future use in separating the free versus protein-bound fractions of small drugs or hormones in biological samples, as directly indicated by the results of this report 7-11. Another expected application would be the use of IA-RAM columns for the separation of labeled/modified peptides from digests or samples containing intact proteins with the same modification or label 49,50…”

Section: Discussionmentioning

confidence: 88%

Development of Immunoaffinity Restricted Access Media for Rapid Extractions of Low-Mass Analytes

Mallik

Hage

2008

Anal. Chem.

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Restricted access media using antibodies as immobilized ligands were developed for the rapid and selective capture of small analytes by immunoextraction, giving rise to materials referred to as immunoaffinity restricted access media (IA-RAM). To make such a material, intact antibodies for the desired target were first immobilized onto porous silica, with antibodies at or near the outer surface of the support then being treated with papain (or a related agent) to release and remove their binding domains. The result was a support in which only antibodies deep within the pores remained intact and able to bind to the target. Items evaluated in the development of such media included the immobilization method used for the antibodies, the pore size of the support, and the amount of papain and time that were used for support treatment. A theoretical model was also developed to describe the extent of binding domain removal based on the measured polypeptide content of the IA-RAM support before and after treatment with papain. The final optimized conditions for making the IA-RAM supports were used to prepare columns that contained anti-fluorescein antibodies. Injections of fluorescein and fluorescein-labeled bovine serum albumin onto these IA-RAM columns gave selective and quantitative extraction of fluorescein in 1-2 s. This approach can be used with other antibodies and low mass targets and should be valuable for such applications as the rapid separation of drugs from drug-protein complexes or the isolation of labeled/modified peptides from intact proteins that contain the same modification or label.

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“…Amidated conjugates of (R)-and (S)-ibuprofen with glycine (2a and 2b) and taurine (3a and 3b), as well as p-nitorophenyl esters of (R)-and (S)-ibuprofen (4a and 4b) were synthesized in our laboratory by a previously reported method [20] (Fig. 1).…”

Section: Methodsmentioning

confidence: 99%

Enantioselective immunorecognition of protein modification with optically active ibuprofen using polyclonal antibody

Ito

Ishiwata

Kosaka

et al. 2004

Journal of Chromatography B

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The Enantioselective Immunoaffinity Extraction of an Optically Active Ibuprofen-Modified Peptide Fragment

Cited by 14 publications

References 32 publications

Effect of the mobile phase on antibody-based enantiomer separations of amino acids in high-performance liquid chromatography

Effect of the mobile phase on antibody-based enantiomer separations of amino acids in high-performance liquid chromatography

Development of Immunoaffinity Restricted Access Media for Rapid Extractions of Low-Mass Analytes

Enantioselective immunorecognition of protein modification with optically active ibuprofen using polyclonal antibody

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