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New biomedical devices with selective peptide recognition properties. Part 1: Characterization and cytotoxicity of molecularly imprinted polymers.

Rechichi A, Cristallini C, Vitale U, Ciardelli G, Barbani N, Vozzi G, Giusti P - J. Cell. Mol. Med. (2007 Nov-Dec)

Bottom Line: The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones).The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour.The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical Engineering, Politecnico di Torino, Torino, Italy.

ABSTRACT
Molecular imprinting is a technique for the synthesis of polymers capable to bind target molecules selectively. The imprinting of large proteins, such as cell adhesion proteins or cell receptors, opens the way to important and innovative biomedical applications. However, such molecules can incur into important conformational changes during the preparation of the imprinted polymer impairing the specificity of the recognition cavities. The "epitope approach" can overcome this limit by adopting, as template, a short peptide sequence representative of an accessible fragment of a larger protein. The resulting imprinted polymer can recognize both the template and the whole molecule thanks to the specific cavities for the epitope. In this work two molecularly imprinted polymer formulations (a macroporous monolith and nanospheres) were obtained using the protected peptide Z-Thr-Ala-Ala-OMe, as template, and Z-Thr-Ile-Leu-OMe, as analogue for the selectivity evaluation, methacrylic acid, as functional monomer, and trimethylolpropane trimethacrylate and pentaerythritol triacrylate (PETRA), as cross-linkers. Polymers were synthesized by precipitation polymerization and characterized by standard techniques. Polymerization and rebinding solutions were analyzed by high performance liquid chromatography. The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones). The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour. The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities. The cytotoxicity tests showed normal adhesion and proliferation of fibroblasts cultured in the medium that was put in contact with the imprinted polymers.

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Related in: MedlinePlus

FT-IR superimposed spectra of TAA peptide (dotted line) and a couple of imprinted and related control polymers. CP 1− (continuous line) and MIP 1− (dashed line) spectra are identical in spite of the presence of the peptide in MIP 1−.
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fig04: FT-IR superimposed spectra of TAA peptide (dotted line) and a couple of imprinted and related control polymers. CP 1− (continuous line) and MIP 1− (dashed line) spectra are identical in spite of the presence of the peptide in MIP 1−.

Mentions: FT-IR spectra showed the characteristic absorption peaks of the expected chemical structure (ν O H = 3450 cm−1; ν C-H = 2970−1cm; ν C = O = 1730 cm−1; ν C(= O)−O = 1160 cm−1). When a lower cross-linker amount was used (for example CP 1− with respect to CP 1+) a more intense band at 3450/cm due to the O-H stretching of the MAA carboxyl group was observed (Fig. 2). When PETRA was used instead of TRIM as cross-linker agent (Fig. 3) a stronger ν O-H band, due to the addition of the side chain C-OH group absorption, and a peak at 1065 cm−1, due to the ν C-O, were noted. The comparison between MIPs and CPs did not highlight the presence of the target molecule, because of the small amount (<3%) of TAA entrapped by the polymers. The spectra of CP 1− and MIP 1−, in Figure 4, do not show any difference in the region around (1550 cm−1) where the target molecule presents an intense absorption peak.


New biomedical devices with selective peptide recognition properties. Part 1: Characterization and cytotoxicity of molecularly imprinted polymers.

Rechichi A, Cristallini C, Vitale U, Ciardelli G, Barbani N, Vozzi G, Giusti P - J. Cell. Mol. Med. (2007 Nov-Dec)

FT-IR superimposed spectra of TAA peptide (dotted line) and a couple of imprinted and related control polymers. CP 1− (continuous line) and MIP 1− (dashed line) spectra are identical in spite of the presence of the peptide in MIP 1−.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4401303&req=5

fig04: FT-IR superimposed spectra of TAA peptide (dotted line) and a couple of imprinted and related control polymers. CP 1− (continuous line) and MIP 1− (dashed line) spectra are identical in spite of the presence of the peptide in MIP 1−.
Mentions: FT-IR spectra showed the characteristic absorption peaks of the expected chemical structure (ν O H = 3450 cm−1; ν C-H = 2970−1cm; ν C = O = 1730 cm−1; ν C(= O)−O = 1160 cm−1). When a lower cross-linker amount was used (for example CP 1− with respect to CP 1+) a more intense band at 3450/cm due to the O-H stretching of the MAA carboxyl group was observed (Fig. 2). When PETRA was used instead of TRIM as cross-linker agent (Fig. 3) a stronger ν O-H band, due to the addition of the side chain C-OH group absorption, and a peak at 1065 cm−1, due to the ν C-O, were noted. The comparison between MIPs and CPs did not highlight the presence of the target molecule, because of the small amount (<3%) of TAA entrapped by the polymers. The spectra of CP 1− and MIP 1−, in Figure 4, do not show any difference in the region around (1550 cm−1) where the target molecule presents an intense absorption peak.

Bottom Line: The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones).The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour.The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical Engineering, Politecnico di Torino, Torino, Italy.

ABSTRACT
Molecular imprinting is a technique for the synthesis of polymers capable to bind target molecules selectively. The imprinting of large proteins, such as cell adhesion proteins or cell receptors, opens the way to important and innovative biomedical applications. However, such molecules can incur into important conformational changes during the preparation of the imprinted polymer impairing the specificity of the recognition cavities. The "epitope approach" can overcome this limit by adopting, as template, a short peptide sequence representative of an accessible fragment of a larger protein. The resulting imprinted polymer can recognize both the template and the whole molecule thanks to the specific cavities for the epitope. In this work two molecularly imprinted polymer formulations (a macroporous monolith and nanospheres) were obtained using the protected peptide Z-Thr-Ala-Ala-OMe, as template, and Z-Thr-Ile-Leu-OMe, as analogue for the selectivity evaluation, methacrylic acid, as functional monomer, and trimethylolpropane trimethacrylate and pentaerythritol triacrylate (PETRA), as cross-linkers. Polymers were synthesized by precipitation polymerization and characterized by standard techniques. Polymerization and rebinding solutions were analyzed by high performance liquid chromatography. The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones). The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour. The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities. The cytotoxicity tests showed normal adhesion and proliferation of fibroblasts cultured in the medium that was put in contact with the imprinted polymers.

Show MeSH
Related in: MedlinePlus