Exploration into the Synthesis and Analysis of a Novel Sensor for Biological Metal Ions
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Exploration into the Synthesis and Analysis of a Novel Sensor for Biological Metal Ions Alexis Kasparian Advisor: Dr. Roy Planalp [email protected]; University of New Hampshire, Parsons Hall, 23 Academic Way, Durham NH 03824 Introduction The synthesis of ligands with particular affinities for metal ions allows their use in sensing. In particular, this research group has worked towards the synthesis of ligands with regard to the trace biological ions zinc(II), copper(II), and iron(II). Upon successful synthesis of ligands and determination of their formation constants with the ions, the ligands may be made functional via an acrylamido group for copolymerization with n-isopropylacrylamide (NIPA). The polyNIPAm system also contains sites that allow for addition of fluorophores. 1 The fluorophores enable FRET under certain conditions, which is used for quantitative detection of metal ions (Figure 2). References [1] Yao, S. et al. Analyst. 2012. 137, 4734-4741. [2] Reddel, J. B.S. Thesis, University of New Hampshire, 2011. [3] Planalp, R.P. et al. Biochem. Soc. Trans. 2002. 30, 758-762 Conclusions and Future Work The computed results show promise in regards to the specificity of the ligand, and progress has been made towards its synthesis. The model reaction has been run under several different conditions and the products are under analysis. Future work for the research group involves: •Successful ligand synthesis •Comparison of its crystal structure, if isolable, to the computed models •Titrations of the ligand to determine its pKa •Titrations of the ligand with iron(II), copper(II), and zinc(II) to determine formation constants •Addition of the ligand onto the polymer delivery system and resultant fluorescent studies with iron(II) concentrations Figure 1. Previously synthesized metal-specific ligands with acrylamido groups for polymerization, left, and general polymer setup, right. 1,2 Link to Current Research To design a sensor selective for iron(II), the previously synthesized tachpyr molecule (Figure 3, left) was examined. Tachpyr is known to favor iron over zinc and is cytotoxic to cells, allowing its use as a cancer cell treatment. 3 To produce a molecule with a reversible binding ability for sensing, tachpyr was modified (Figure 3, right). This novel molecule is currently under study. Synthetic routes are being explored and computational modeling is employed to determine how it will interact with the aforementioned metal ions. Figure 3. Tachpyr molecule and subsequent theoretical modification for desired sensor ligand. Original Synthesis Route One synthesis route was explored (Figure 6), but after several attempts it was determined that the yield of conversion of 2 to 3 was too low for our purposes. The approach of Figure 7 appears more suitable. Figure 8. Model reaction for determination of reaction conditions using 2-aminomethyl pyridine with 1,3-propanediol di-p-tosylate. Figure 6. Original synthesis route. Conversion of 2 to 3 was very low yielding, prompting research into alternate methods for synthesis. NH HN NH N N N N N H N H N -removal of one aminomethyl pyridine group and subsequent loss of cyclohexyl ring -addition ofmethyl groups toprevent imine formation Current Synthesis Routes and Results The synthesis of the desired ligand is in progress (Figure 7). We are working to improve the outcome of the amine formation (3) by rigorous exclusion of water and oxygen, which may quench the formation of the necessary organocerium complex. The displayed plans do not include the acrylamido group for polymerization. Its synthesis could be accomplished with the introduction of a nitro group on the center carbon of 1,3-propanediol di-p-tosylate, which can then be converted to an amine for reaction to the resulting acrylamido moiety. Figure 2. Mechanism of action of the ligand-NIPA copolymer. The ratio of donor to acceptor emission, when FRET occurs or does not occur, is used to determine the amount of metal ion bound to the polymer. TsO OTs HO OH 8 6 Synthesis of 6 : C 5 H 5 N TsCl 62 % N Br N OH N N N H O NH 2 N N H N H H 3 C CH 3 H 3 C CH 3 N TsO OTs <1/2 1.6 M HCl 1.BF 3 Et 2 O, CH 3 CN 1.nBuLi, THF 2.CH 3 COCH 3 3. NH 4 Cl Br Br Br N N H O KOH, MeOH 2.basify 30% NaOH 1 2 3 4 5 6 7 2.NaOH 85 % 8.3 % 51 % Raney nickel H 3 C CH 3 H 3 C CH 3 CH 3 H 3 C H 3 C CH 3 Figure 7. Alternate synthesis route, in progress. Figure 5. Equation used to compare binding of the model ligand with different metal ions. Table 1. Calculated results of relative energies using the equation in Figure 5. Figure 4. Four low energy conformers of the ligand attached to a metal ion center. Two water molecules complete the octahedral geometry. N N H H 3 C CH 3 N H N H 3 C CH 3 N C N N NH 2 H 3 C CH 3 TsO OTs < 1/2 1.CeCl 3 ,THF, -78°C H 3 C Li 2.NH 4 OH 1 2 3 4 5 TsO OTs HO OH 6 4 Synthesis of 4: C 5 H 5 N TsCl 62 % N NH 2 TsO OTs solvent, 2.5 eq base heat/reflux for X hr N N H N H N 2 eq O NH NH O ligand O NH NH 2 x y z NIPA site for fluoropho addition To determine the most practical way for the addition of two amines onto a single di- tosylate, a model reaction is under exploration. This reaction (Figure 8) involves the use of 2-aminomethylpyridine, a readily available reagent. It is being reacted under different conditions to determine which method is most efficient for completion of reaction. N N N O bipy PEPMA N N N HN O The results of the computations show that for Cases 1-3 the equilibrium will prefer the left side of the equation, with iron(II) bound to the ligand. Case 4 shows preference for copper binding, so it is possible that the ligand may have some affinity for copper(II). Acknowledgements Thank you to the UNH Department of Chemistry for their continuing support. My sincerest thanks to Dr. Planalp and Lea Nyiranshuti of UNH for their guidance, and Dr. Richard Johnson for his introduction to computational models. We thank the UNH URA and Craig West Undergraduate Award programs Zn (II) complex Cu(II) complex Fe(II) high spin complex Fe(II) low spin complex M 1 -ligand(H 2 O) 2 + M 2 (H 2 O) 6 M 2 -ligand(H 2 O) 2 + M 1 (H 2 O) 6 Case 1: M 1 = Fe II ,M 2 = Zn II (low spin iron) Case 2: M 1 = Fe II ,M 2 = Cu II (low spin iron) Case 3: M 1 = Fe II ,M 2 = Zn II (high spin iron) Case 4: M 1 = Fe II ,M 2 = Cu II (high spin iron) Total Energy Differences, right side - left side [Hartree] [kcal/ mol] Case 1 0.04433 27.82 Case 2 0.01939 12.17 Case 3 0.01675 10.51 Case 4 -0.00819 -5.14 h h ' h '' increasing temperature h metal ion h h ' AlexaFluor 647 labeled acceptor strand AlexaFluor 555 labeled donor strand h ' donor emission acceptor emission h '' FRET Computational Results The model ligand was analyzed using Spartan software modeling programs. Energy comparisons of the ligand bound to the various ions were performed to determine likely binding results. Lowest energy conformers of the ligand were developed, assuming a tetradentate binding fashion with the ligand and completion of an octahedral geometry with water molecules (Figure 4). The conformers’ energies were then compared (Figure 5, Table 1).
description
Exploration into the Synthesis and Analysis of a Novel Sensor for Biological Metal Ions. Alexis Kaspa rian Advisor: Dr. Roy Planalp [email protected] du; University of New Hampshire, Parsons Hall, 23 Academic Way, Durham NH 03824. Introduction - PowerPoint PPT Presentation
Transcript of Exploration into the Synthesis and Analysis of a Novel Sensor for Biological Metal Ions
Exploration into the Synthesis and Analysis of a Novel Sensor for Biological Metal Ions
Alexis Kasparian Advisor: Dr. Roy [email protected]; University of New Hampshire, Parsons Hall, 23 Academic Way, Durham NH
03824
IntroductionThe synthesis of ligands with particular affinities for metal ions allows their use in sensing. In particular, this research group has worked towards the synthesis of ligands with regard to the trace biological ions zinc(II), copper(II), and iron(II). Upon successful synthesis of ligands and determination of their formation constants with the ions, the ligands may be made functional via an acrylamido group for copolymerization with n-isopropylacrylamide (NIPA). The polyNIPAm system also contains sites that allow for addition of fluorophores.1 The fluorophores enable FRET under certain conditions, which is used for quantitative detection of metal ions (Figure 2).
References [1] Yao, S. et al. Analyst. 2012. 137, 4734-4741.[2] Reddel, J. B.S. Thesis, University of New Hampshire,
2011.[3] Planalp, R.P. et al. Biochem. Soc. Trans. 2002. 30,
758-762
Conclusions and Future WorkThe computed results show promise in regards to the specificity of the ligand, and progress has been made towards its synthesis. The model reaction has been run under several different conditions and the products are under analysis.
Future work for the research group involves:•Successful ligand synthesis•Comparison of its crystal structure, if isolable, to the computed models•Titrations of the ligand to determine its pKa•Titrations of the ligand with iron(II), copper(II), and zinc(II) to determine formation constants•Addition of the ligand onto the polymer delivery system and resultant fluorescent studies with iron(II) concentrations
Figure 1. Previously synthesized metal-specific ligands with acrylamido groups for polymerization, left, and general polymer setup, right.1,2
Link to Current ResearchTo design a sensor selective for iron(II), the previously synthesized tachpyr molecule (Figure 3, left) was examined. Tachpyr is known to favor iron over zinc and is cytotoxic to cells, allowing its use as a cancer cell treatment.3 To produce a molecule with a reversible binding ability for sensing, tachpyr was modified (Figure 3, right). This novel molecule is currently under study. Synthetic routes are being explored and computational modeling is employed to determine how it will interact with the aforementioned metal ions.
Figure 3. Tachpyr molecule and subsequent theoretical modification for desired sensor ligand.
Original Synthesis RouteOne synthesis route was explored (Figure 6), but after several attempts it was determined that the yield of conversion of 2 to 3 was too low for our purposes. The approach of Figure 7 appears more suitable.
Figure 8. Model reaction for determination of reaction conditions using 2-aminomethyl pyridine with 1,3-propanediol di-p-tosylate.
Figure 6. Original synthesis route. Conversion of 2 to 3 was very low yielding, prompting research into alternate methods for synthesis.
NH
HN NHN N
NN
NH
NH
N
-removal of one aminomethyl pyridine group and subsequent loss of cyclohexyl ring-addition of methyl groups to prevent imine formation
Current Synthesis Routes and ResultsThe synthesis of the desired ligand is in progress (Figure 7). We are working to improve the outcome of the amine formation (3) by rigorous exclusion of water and oxygen, which may quench the formation of the necessary organocerium complex.
The displayed plans do not include the acrylamido group for polymerization. Its synthesis could be accomplished with the introduction of a nitro group on the center carbon of 1,3-propanediol di-p-tosylate, which can then be converted to an amine for reaction to the resulting acrylamido moiety.
Figure 2. Mechanism of action of the ligand-NIPA copolymer. The ratio of donor to acceptor emission, when FRET occurs or does not occur, is used to determine the amount of metal ion bound to the polymer.
TsO OTsHO OH
8 6
Synthesis of 6:
C5H5N
TsCl
62 %
N
Br
N
OH
N
N
NH
O
NH2
N
NH
NH
H3C CH3H3C CH3
N
TsO OTs<1/2
1. 6 M HCl
1. BF3 Et2O, CH3CN1. nBuLi, THF
2. CH3COCH3
3. NH4Cl
Br Br Br
N
NH
O
KOH, MeOH
2. basify 30% NaOH
1 2 3
4
5
6
7
2. NaOH
85 %8.3 % 51 %
Raney nickel
H3C CH3 H3C CH3 CH3H3C
H3C CH3
Figure 7. Alternate synthesis route, in progress.
Figure 5. Equation used to compare binding of the model ligand with different metal ions.
Table 1. Calculated results of relative energies using the equation in Figure 5.
Figure 4. Four low energy conformers of the ligand attached to a metal ion center. Two water molecules complete the octahedral geometry. N
NH
H3C CH3
NH
N
H3C CH3
N
CN
N
NH2
H3C CH3
TsO OTs< 1/2
1. CeCl3, THF, -78 °CH3C Li
2. NH4OH
1
2
3
4
5
TsO OTsHO OH6 4
Synthesis of 4:
C5H5NTsCl
62 %
N
NH2TsO OTs
solvent, 2.5 eq baseheat/reflux for X hr
N
NH
NH
N2 eq
O NH NHO
ligand
O NH
NH2
x y z
NIPA
site for fluorophore addition
To determine the most practical way for the addition of two amines onto a single di-tosylate, a model reaction is under exploration. This reaction (Figure 8) involves the use of 2-aminomethylpyridine, a readily available reagent. It is being reacted under different conditions to determine which method is most efficient for completion of reaction.
N
N
N
O
bipyPEPMAN
N
N
HN
O
The results of the computations show that for Cases 1-3 the equilibrium will prefer the left side of the equation, with iron(II) bound to the ligand. Case 4 shows preference for copper binding, so it is possible that the ligand may have some affinity for copper(II).
Acknowledgements
Thank you to the UNH Department of Chemistry for their continuing support. My sincerest thanks to Dr. Planalp and Lea Nyiranshuti of UNH for their guidance, and Dr. Richard Johnson for his introduction to computational models. We thank the UNH URA and Craig West Undergraduate Award programs for funding.
Zn (II) complex Cu(II) complex
Fe(II) high spin complex
Fe(II) low spin complex
M1-ligand(H2O)2 + M2(H2O)6 M2-ligand(H2O)2 + M1(H2O)6
Case 1: M1 = FeII, M2 = ZnII (low spin iron)
Case 2: M1 = FeII, M2 = CuII (low spin iron)
Case 3: M1 = FeII, M2 = ZnII (high spin iron)
Case 4: M1 = FeII, M2 = CuII (high spin iron)
Total Energy Differences, right side - left side[Hartree] [kcal/mol]
Case 1 0.04433 27.82Case 2 0.01939 12.17Case 3 0.01675 10.51Case 4 -0.00819 -5.14
h
h'
h''
increasing temperature
h
metal ion
h
h'
AlexaFluor 647 labeled acceptor strand
AlexaFluor 555 labeled donor strand h' donor emission
acceptor emissionh''
FRET
Computational ResultsThe model ligand was analyzed using Spartan software modeling programs. Energy comparisons of the ligand bound to the various ions were performed to determine likely binding results. Lowest energy conformers of the ligand were developed, assuming a tetradentate binding fashion with the ligand and completion of an octahedral geometry with water molecules (Figure 4). The conformers’ energies were then compared (Figure 5, Table 1).
