The Cation Specific Effects on the Aqueous Solubility of Amino Acids

85 Pages·2014·2.81 MB·English

The Cation Specific Effects on the Aqueous Solubility of Amino Acids:
Experimental and Molecular Dynamics Simulations Contributions

Cátia Sofia Ribeiro De Sousa

Escola Superior de Tecnologia e de Gestão
Instituto Politécnico de Bragança

Master's degree in chemical engineering

July 2014
The Cation Specific Effects on the Aqueous Solubility of Amino
Acids: Experimental and Molecular Dynamics Simulations
Contributions

Cátia Sofia Ribeiro De Sousa

Escola Superior de Tecnologia e de Gestão
Instituto Politécnico de Bragança

Master's degree in chemical engineering

Supervisors:
Dr. Luciana Tomé
Professor Olga Ferreira
Professor Simão Pinho

July 2014
To my Father.
iii
Acknowledgements
I would like to express my gratitude to my supervisors Professor Simão Pinho and Professor Olga
Ferreira for the opportunity to integrate this research project, the possibility of collaboration with
CICECO Laboratory at the University of Aveiro, the scientific guidance, and the overall support
during the research project.
This work would not be possible without the help of Dr. Luciana Tomé. I am grateful for the
scientific guidance, and for her patience and kindness that she showed guiding my research
activities.
My grateful acknowledge to Escola Superior de Tecnologia e Gestão (ESTiG) directed by
Professor Albano Alves for providing the laboratory space and equipment needed.
I am very grateful to Professor João Coutinho for providing the laboratory space and equipment
needed, as well as his support during the research project.
I thank my Path and LSRE/IPB colleagues for the companionship and cooperation.
Finally, special thanks to all those who have been part of my life, for all the affection, dedication
and encouragement. Thanks to my Mother and to Lino Maia.

iv
Resumo
O estudo dos efeitos específicos dos iões na solubilidade de aminoácidos e proteínas, em
soluções aquosas, é vital para o desenvolvimento de muitas áreas da bioquímica e da
biotecnologia.
Neste trabalho estudou-se o efeito dos iões de sais inorgânicos na solubilidade de
aminoácidos, em soluções aquosas salinas, através de medições experimentais e simulações de
dinâmica molecular. Foram selecionados três aminoácidos (DL-alanina, L-isoleucina e L-valina)
como compostos modelo. Face à sua relevância biológica e escassa informação disponível na
literatura, selecionaram-se ainda os seguintes catiões: K+, Li+, Ca2+ e Al3+, variando o anião entre
Cl- ou .

Recorrendo a um método analítico isotérmico mediu-se a solubilidade, a 298,15 K, da
DL-alanina (Ala) em soluções aquosas contendo CaCl e K SO e da L-isoleucina (Iso) e L-
2 2 4
valina (Val), em soluções aquosas contendo CaCl , K SO , LiCl, Li SO , Al (SO ) e AlCl . No
2 2 4 2 4 2 4 3 3
caso de LiCl, o efeito observado na solubilidade de Val e Iso é praticamente inexistente enquanto
Li SO e K SO induzem um efeito de salting-out. Por último, para CaCl , Al (SO ) e AlCl
2 4 2 4 2 2 4 3 3
observou-se um efeito de salting-in bastante pronunciado, sendo mais forte no caso dos sais que
contêm alumínio.
Os resultados obtidos pelas simulações de dinâmica molecular sugerem que o efeito de
salting-in na solubilidade é diferente entre catiões e aniões. Os catiões polivalentes fortemente
hidratados (Ca2+ e Al3+) apresentam interações extremamente fortes com o grupo carboxílico dos
aminoácidos, resultando num efeito pronunciado de salting-in. Por outro lado, os iões Li+ e K+
não estabelecem interações relevantes com os aminoácidos, apresentando pouco efeito na
solubilidade.

Palavras-chave: aminoácidos, solubilidade, efeito de iões, dinâmica molecular

v
Abstract
The study of ion specific effects on the aqueous solubility of amino acids and proteins is
crucial for the development of many areas of biochemistry and biotechnology.
In this work, the effect of ions of inorganic salts on the aqueous solubility of amino acids
was studied by performing experimental solubility measurements and molecular dynamics
simulations. Three amino acids (DL-alanine, L-isoleucine and L-valine) were chosen as model
compounds. Taking into consideration their biological relevance and scarce available
information, cations such as K+, Li+, Ca2+and Al3+ were selected, varying the anion between Cl-
or .

New solubility data were measured, at 298.15 K, using the analytical isothermal shake-
flask method, for DL-alanine (Ala) in aqueous solutions of CaCl and K SO , and for L-
2 2 4
isoleucine (Ile) and L-valine (Val) in aqueous solutions of CaCl , K SO , LiCl, Li SO , Al (SO )
2 2 4 2 4 2 4 3
and AlCl . For LiCl no significant solubility effect was observed for Val and Ile while Li SO
3 2 4
and K SO induced a salting-out effect. In the case of CaCl , Al (SO ) and AlCl , a pronounced
2 4 2 2 4 3 3
salting-in was observed, much stronger for the aluminum salts.
The molecular dynamics simulations results obtained suggest that the solubility effects of
salting-in inducing cations is different from that of the anions. Strongly hydrated polyvalent
cations (Ca2+ and Al3+) present extremely strong interactions with the carboxylic group of the
amino acids, resulting in a pronounced salting-in effect. On the other hand, Li+ and K+ ions do
not establish important interactions with the amino acids, having thus a small effect on their
aqueous solubility.

Keywords: amino acids, solubility, ion effects, molecular dynamics

vi
Table of Contents
List of Figures ................................................................................................................................. ix
List of Tables .................................................................................................................................. xv
Chapter 1. Introduction ................................................................................................................ 1
1.1. Motivation and objectives ................................................................................................. 1
1.2. Structure of the thesis ........................................................................................................ 1
1.3. State of the art .................................................................................................................... 2
1.3.1. The salt effect on the solubility of amino acids and proteins in water ....................... 2
1.3.2. Experimental solubility database ............................................................................... 5
Chapter 2. Experimental part ..................................................................................................... 13
2.1. Materials and Methods .................................................................................................... 13
2.1.1. Chemical compounds ............................................................................................... 13
2.1.2. Experimental procedure ........................................................................................... 14
2.2. Results and discussion ..................................................................................................... 17
2.2.1. Solubility in water .................................................................................................... 17
2.2.2. Solubility in aqueous saline solutions ...................................................................... 17
2.2.3. pH measurements ..................................................................................................... 22
Chapter 3. Molecular Dynamics Simulation .............................................................................. 23
3.1. Introduction ..................................................................................................................... 23
3.2. Computational method .................................................................................................... 25
3.3. Results and discussion ..................................................................................................... 26
3.3.1. Anion effect .............................................................................................................. 28
3.3.2. Cation effect ............................................................................................................. 33
3.3.3. Interactions with water ............................................................................................. 42
3.3.4. Amino acid side chain effect .................................................................................... 45
vii
3.3.4.1. Anions ................................................................................................................... 45
3.3.4.2. Cations .................................................................................................................. 46
Chapter 4. Conclusions and Future Work .................................................................................. 49
References ...................................................................................................................................... 51
Appendix A .................................................................................................................................... 57

viii
List of Figures
Figure 1 - Typical order of cations and anions in the Hofmeister series (adapted from Kunz,
2010). ................................................................................................................................................ 2
Figure 2 - Relative solubility of DL-alanine in aqueous saline solutions, at 298.15 K, containing:
◊, NaF (El-Dossoki 2010); ■, KF (El-Dossoki 2010). ..................................................................... 6
Figure 3 - Relative solubility of DL-alanine in aqueous saline solutions, at 298.15 K, containing:
♦, NaBr (El-Dossoki 2010); +, KBr (El-Dossoki 2010). .................................................................. 6
Figure 4 - Relative solubility of DL-alanine in aqueous saline solutions, at 298.15 K, containing:
▲, K SO (El-Dossoki 2010); Δ, (NH ) SO (Ferreira et al. 2009). ............................................... 7
2 4 4 2 4
Figure 5 - Relative solubility of DL-alanine in aqueous saline solutions, at 298.15 K, containing:
Δ, KCl (Khoshkbarchi and Vera 1997); ○, KCl (Ferreira et al. 2005); □, CaCl (El-Dossoki
2
2010); □, NH Cl (Tomé et al. 2013); ×, NaCl (Khoshkbarchi and Vera 1997); ●, BaCl (El-
4 2
Dossoki 2010). ................................................................................................................................. 7
Figure 6 - Relative solubility of L-valine in aqueous solutions, at 298.15 K, containing: ◊, NaF
(El-Dossoki 2010); ■, KF (El-Dossoki 2010). ................................................................................. 8
Figure 7 - Relative solubility of L-valine in aqueous solutions, at 298.15 K, containing: ♦, NaBr
(El-Dossoki 2010); +, KBr (El-Dossoki 2010). ............................................................................... 8
Figure 8 - Relative solubility of L-valine in aqueous solutions containing, at 298.15 K,: □, CaCl
2
(El-Dossoki 2010); ●, BaCl (El-Dossoki 2010); □, NH Cl (Tomé et al. 2013); ○, MgCl (Tomé
2 4 2
et al. 2013). ....................................................................................................................................... 9
Figure 9 - Relative solubility of L-valine in aqueous solutions, at 298.15 K, containing: ▲,
K SO (El-Dossoki 2010); ▲, (NH ) SO (Tomé et al. 2013); ◊, MgSO (Tomé et al. 2013). ...... 9
2 4 4 2 4 4
Figure 10 - Relative solubility of L-valine in aqueous solutions, at 298.15 K, containing: ×,
Na SO (El-Dossoki 2010); ×, Na SO (Ramasami 2002); ×, Na SO (Islam and Wadi 2001). ... 10
2 4 2 4 2 4
Figure 11 - Relative solubility of L-isoleucine in aqueous solutions, at 298.15 K, containing: Δ,
(NH ) SO (Ferreira et al. 2009); ◊,MgSO (Tomé et al. 2013). ................................................... 10
4 2 4 4
Figure 12 - Relative solubility of L-isoleucine in aqueous solutions, at 298.15 K, containing: □,
KCl (Ferreira et al. 2007); □, NH Cl (Tomé et al. 2013); ○, MgCl (Tomé et al. 2013). ............. 11
4 2
Figure 13 - Structure of the amino acids studied in this work: (a) alanine, (b) valine and (c)
isoleucine. ....................................................................................................................................... 13
ix
Figure 14 - Experimental setup used on the shake flask method. .................................................. 14
Figure 15 - Density meter used in this work. ................................................................................. 15
Figure 16 - Typical refractive index versus concentration of amino acid curve, obtained by
applying method B. ........................................................................................................................ 16
Figure 17 – Refractometer used in this work. ................................................................................ 16
Figure 18 - Relative solubility of alanine (blue), valine (red), and isoleucine (green) in aqueous
solutions, at 298.15 K, containing the salt: (a) □, LiCl; Δ, CaCl ; and in (b) ◊, Li SO ; ○, K SO .
2 2 4 2 4
Lines are guides to the eyes. ........................................................................................................... 20
Figure 19 - Relative solubility of DL-alanine in potassium sulfate aqueous solutions obtained in
this work and by El-Dossoki (2010), at 298.15 K. ......................................................................... 21
Figure 20 - Relative solubility of DL-alanine in CaCl aqueous solutions obtained in this work
2
and by El-Dossoki (2010), at 298.15 K. ......................................................................................... 21
Figure 21 - Structure and atom labeling of the amino acids studied in this work: (a) alanine (Ala),
(b) valine (Val) and (c) isoleucine (Ile). C stands for the terminal carbon atom of the amino acid
t
side chain and C is used to denote the carbon backbone of the amino acid side chain. ............... 27
B
Figure 22 - Radial distribution functions corresponding to the interactions of isoleucine C with
t
the central atom of the anions (Cl or S), for salts containing Li+ and Al3+ cations. ....................... 28
Figure 23 - Radial distribution functions corresponding to the interactions of isoleucine C with
B
the central atom of the anions (Cl or S), for salts containing Li+ and Al3+ cations. ....................... 29
Figure 24 - Radial distribution functions corresponding to the interactions of the H atom of the
amine group of isoleucine with the central atom of the anions (Cl or S), for salts containing Li+
and Al3+ cations. ............................................................................................................................. 29
Figure 25 - Radial distribution functions corresponding to the interactions of the O atom of the
carboxylic group of isoleucine with the central atom of the anions (Cl or S), for salts containing
Li+ and Al3+ cations. ....................................................................................................................... 30
Figure 26 - Radial distribution functions corresponding to the interactions of Li SO salts with
2 4
water molecules, in the presence of isoleucine. ............................................................................. 33
Figure 27 - Radial distribution functions corresponding to the interactions of isoleucine C with
t
the cations (Li+, K+, Ca2+ or Al3+) for all the studied salts. ............................................................ 34
Figure 28 - Radial distribution functions corresponding to the interactions of isoleucine C with
B
the cations (Li+, K+, Ca2+ or Al3+) for all the studied salts. ............................................................ 34
x