The Effect of Amino Acids on the Solubility of Copper (II) CitraconateRhomesia Ramkellowan

Mentor: Sabrina G. Sobel, Department of Chemistry, Hofstra University, Hempstead, New York, 11549

The Effect of Amino Acids on the Solubility of Copper (II) CitraconateRhomesia Ramkellowan

Mentor: Sabrina G. Sobel, Department of Chemistry, Hofstra University, Hempstead, New York, 11549

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Citraconic acid, copper (II) carbonate and all amino acids were used as received. Na2EDTA was dried for >1 hr at 100°C before use. Preparation of Copper (II) CitraconatePreparation of Copper (II) CitraconateCCitraconic acid was stirred in 20 mL Nanopure H2O until completely dissolved. Copper (II) carbonate was stirred in 20 mL Nanopure H2O on medium heat. Citraconic acid solution was added to stirred copper (II) carbonate solution by pipette. Mixture was left to stir overnight and filtered. Filtrate was placed in rotary evaporator and the solids were dried in a desiccator to yield potentially copper (II) citraconate. Preparation of SolutionsPreparation of Solutions One gram copper (II) citraconate was combined with varying molar amounts of amino acid and 20 mL Nanopure H2O, stirred overnight and diluted to 25 mL. Mixtures were quantitatively filtered after 48 hrs.Analysis of Total Copper(II) Ion ContentAnalysis of Total Copper(II) Ion Content Small aliquots of filtered solution (~2 mL) were combined with 25.0 mL pH 5.5 acetate buffer, and titrated with 0.0100 M Na2EDTA to endpoint using Xylenol Orange indicator.

Calculation of Percent Dissolved Copper(II) Citraconate Calculation of Percent Dissolved Copper(II) Citraconate All titration results were entered into an Excel spreadsheet. Then, based on the stoichiometry of 1:1 Cu2+:EDTA2-, moles copper, and grams copper (II) citraconate, percent copper (II) citraconate dissolved was calculated.

Abstract

Procedure

Copper (II) Citraconate + Amino Acids Relevant Equilibria

Bibliography

UV-Vis Spectra with Added Glycine

Introduction

Determination of interaction of strong and weak complexing agents and effect on free copper(II) ion content can lead to useful insight for development of applications

• Copper(II) complexes are important in biological and industrial contexts– General Nutrition (RDA 900mcg)– Wound healing (promotes debridement and healing of burn wounds)– Plant nutrition (provides an essential nutrient)

• What form of copper(II) is best for each situation?– Are free copper(II) ions (weakly complexed) or strongly complexed copper(II) ions more

beneficial?– Strong complexes for the removal of copper(II) ; weak complexes for the delivery of

copper(II). • Copper(II) complex characteristics

– Strong complexing agents (e.g. citraconate) and weak complexing agents (e.g. amino acids) are well known.

– The interaction of the strong and weak complexing agents and effect on free copper(II) ion content is not known.

– Initial research with zinc gluconate + glycine showed a modest increase in free zinc ion content (93% vs.91%). Will copper(II) complexes act the same?

– Previous research in this group has shown an increase of zinc citrate solubility with the addition of amino acids (w/glycine, 0 - 50 molar excess: range of 10% - 50% soluble)

• Attempted synthesis of copper (II) citraconate was mostly successful• Composition of copper(II) citraconate is still unclear

• Significant initial solubility of copper(II) citraconate (0.6454g/25 mL, or 0.0258 g per mL)• Percent copper(II) citraconate dissolved increased as amino acid ratio increased• Added amino acid dramatically enhances solubility of copper (II) citraconate prepared• Speciation diagrams of citraconic acid and glycine were calculated using known equilibrium constants (Table above)• Enhancement of solubility cannot solely be due to protonation of salt anion by free amino acid as pK’s of anions and amino acids are comparable

Equilibrium pK Symbol

H2citr H+ + Hcitr- (HL) 2.23 Kcitr1

Hcitr- H+ + citr-2 (L) 5.46 Kcitr2

H2gly+ H+ + Hgly (zwitterion) (HL’) 2.34 Kgly2

Hgly H+ + gly- (L’) 9.60 Kgly3

logK

Cu+2 + HL ZnHL to be determined KHL

Cu+2 + L ZnL 3.4 KL1

Cu+2 + HL’ ZnHL’ 0.12 KHL’

Cu+2 + L’ ZnL’ 4.90† KL’1

CuL’ + L’ ZnL’2 9.01† KL’2

Other Possible Equilibria

(citr gly) Ligand Exchange

CuL + L’ CuL’ + L 1.5†*

CuHL + HL’ CuHL’ + HL to be determined

CuL + HL’ CuHL’ + L to be determined

Glycine Addition to Copper(II) Citraconate

CuHL + HL’ Cu(HL)HL’ to be determined

CuL + HL’ Cu(L)HL’ to be determined

CuL + L’ CuLL’ to be determined

Future Work• Create new method to synthesize pure zinc citraconate and copper (II) citraconate.• Analysis of salt will be done before titrations with amino acids. • Complete trials and UV-Vis analyses with other amino acids. • Expect percent zinc dissolved and percent copper dissolved to increase as amino acid ratio increase.• Measure free [Cu2+] in all solutions using ion specific electrode• Expect to see an increase in free [Cu2+] as the amino acid : zinc salt ratio increases.

Thanks to:Allison Haigney: additional mentor, Lois and Mike: helpful co-researchers, Hofstra U. Chemistry Department: site sponsor,. Dr. Sat Bhattacharya and Harlem Children Society, Ms. Susan Fahrenholtz, Project Seed and the American Chemical Society.

Conclusion

† may not be important at pH 5.5, [L’] = 0.0132% of total [glycine]*not important: K < 1

Zinc and copper salts are known to be important in general human nutrition and surface wound healing. The success formulations that provide zinc and copper(II) ions locally at the wound site may depend on the bioavailability of zinc and copper(II) ions. A study of the influence that amino acids have on the solubility of zinc citraconate may be relevant to such therapy. In this research, copper(II) citraconate was combined with varying molar amounts of amino acids (glycine, alanine and serine) and each solution was titrated with Na2EDTA to determine the total copper(II) ion content in the solutions, and therefore the percent of zinc citraconate that dissolved and the percent copper(II) titrated. It is hoped that this research may help aid future development of medical copper(II) salt treatments.

Serine

Organic Molecules Used

AlanineGlycine

• Copper dissolved starts at ~26% without added glycine.

• Added glycine gradually increases the solubility of Copper (II) citraconate.

• Maximum solubility of ~65% was seen at a glycine to total copper molar ratio of 5:1

• Added serine increases the solubility of Copper (II) Citraconate.

• Maximum solubility of ~61% was seen at a serine to total copper molar ratio of 5:1

•Added alanine gradually increases the solubility of Copper (II) Citraconate.

• Maximum solubility of ~60% was seen at a alanine to copper salt molar ratio of 5:1

Alanine is less polar than glycine, and serine is

more polar that glycine

Citraconic acid

Partially funded by Godfrey Science & Design, Inc., Huntingdon Valley, PA 2006

Project SEED

Copper(II) Citraconate + Glycine

y = -0.0093x2 + 0.122x + 0.2709

R2 = 0.9675

20%

30%

40%

50%

60%

70%

80%

0 2 4 6 8 10Rel. Moles of Glycine

% Cu(Citr) Dissolved

Copper(II) Citraconate + Alanine

y = -0.0076x2 + 0.0976x + 0.3464

R2 = 0.6993

20%

30%

40%

50%

60%

70%

0 2 4 6 8 10 12Rel. Moles of Alanine

% CuCitr Dissolved

Peak Maxima ShouldersCu(Citr):Glycine Ratio Wavelength (nm) Absorbance Wavelength (nm) Absorbance

1:0 750 0.290 none

1:1 742 0.402 640-665 0.20-0.24

1:2 736 0.539 630-665 0.27-0.30

1:5 724 0.617 600-665 0.26-0.36

Copper Citraconate Plus Glycine

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

400 500 600 700 800 900

Wavelength (nm)

Absorbance

1:0

1:1

1:2

1:5

Copper(II) Citraconate + Serine

y = 0.0557x + 0.3588

R2 = 0.6372

20%

30%

40%

50%

60%

70%

0 1 2 3 4 5 6

Rel. Moles Serine

% CuCitr Dissolved