Synthesis and physico-chemical characterization of fatty esters
with an aromatic part.
P. P. P.
P. SainvituSainvituSainvituSainvitua,ba,ba,ba,b, , , , K. Nott K. Nott K. Nott K. Nott a,ca,ca,ca,c,,,, F. NicksF. NicksF. NicksF. Nicksa,ca,ca,ca,c, P. Laurent, P. Laurent, P. Laurent, P. Laurenta,ca,ca,ca,c, L. , L. Willems, L. , L. WillemsWillemsWillemsdddd, J., J.----P. , J., J.P. P. P. CosseCosseCosseCossedddd, , , ,
J. J. J.
J.----M. M. M. CrowetM. CrowetCrowetCrowetbbbb, , , , LLLL. . . . LinsLinsLinsLinsbbbb, , , , C. C. JérômeC. C. JérômeJérômeJérômeeeee, , , , M. M. DeleuM. M. DeleuDeleuDeleua,ba,ba,ba,b....
a Department of Industrial and Biological Chemistry, bCentre de Biophysique Moléculaire Numérique, c Department of General and Organic Chemistry, dMolecular and Cellular Biology lab, Gembloux Agro-Bio Tech, University of Liege, Belgium, eCenter for Education and Research on Macromolecules, University of Liege, Belgium
We thank the Belgian French Community – Concerted Research Actions – Wallonia - Europe Academy for financial support. MD and LL thank the FNRS for their position as research associate and senior research associate ,
respectively. FN and PL are working for the Technose project (Walloon Region).
Louvain-la-Neuve, Nov 16th 2012
Acknowledgments
2222
N DN DN D S T EPN D S T EPS T EPS T EP: B
: B
: B
: B
I OPHYSICALI OPHYSICALI OPHYSICALI OPHYSICALC
C
C
C
HARACTERIZATIONHARACTERIZATIONHARACTERIZATIONHARACTERIZATIONInterfacial properties
C
C
C
C
ONCLUSIONONCLUSIONONCLUSIONONCLUSION TheTheThe
The additionadditionadditionaddition ofofof aaaa fattyof fattyfattyfatty chainchainchainchain onon aromaticononaromaticaromaticaromatic moleculesmoleculesmoleculesmolecules enhancesenhancesenhancesenhances theirtheirtheirtheir interfacialinterfacial propertiesinterfacialinterfacialpropertiespropertiesproperties.... AllAllAllAll thethethethe testedtestedtested moleculestestedmolecules showedmoleculesmoleculesshowedshowedshowed anananan affinityaffinity foraffinityaffinityforforfor thethethethe air/water
air/waterair/water
air/water interfaceinterfaceinterfaceinterface.... ItItItIt increasedincreasedincreasedincreased withwith substitutionwithwithsubstitutionsubstitutionsubstitution ofofofof thethethethe aromaticaromaticaromaticaromatic ringring withringringwithwithwith OHOHOH andOHandand OMeandOMeOMe groupsOMegroups.... Theygroupsgroups TheyTheyThey spontaneouslyspontaneouslyspontaneously penetratespontaneouslypenetratepenetratepenetrate membranesmembranesmembranesmembranes.... Their
TheirTheir
Their interactioninteraction withinteractioninteractionwithwith thewiththethe membranethemembranemembranemembrane couldcouldcouldcould bebe atbebeatatat thethethethe originoriginoriginorigin ofofofof cellcell toxicitycellcelltoxicitytoxicity duetoxicitydueduedue tototo aaaa phenomenonto phenomenonphenomenonphenomenon ofofof membraneofmembrane destabilizationmembranemembranedestabilizationdestabilizationdestabilization.... TheTheThe toxicityThetoxicitytoxicitytoxicity ofofofof phenolic
phenolicphenolic
phenolic fattyfattyfatty estersfattyestersestersesters isisisis concentrationconcentrationconcentrationconcentration----dependentdependent.... Independentdependent InInIn thethethethe future,future,future,future, theirtheirtheirtheirin----vivoinininvivovivovivo antioxidantantioxidantantioxidantantioxidant abilityabilityabilityability atatat lowatlow----concentrationlowlowconcentrationconcentration willconcentrationwillwillwill bebebebe studied,studied,studied,studied,
In Silico
prediction of Interactions with membranes
Fig 1. Compression isotherms recorded with a Langmuir film balance (KSV, Finland). Spreading of pure compounds (25µl, 2mM) on milliQ water at 25°C. The aromatic reagents were not able to form stable monolayers.
Fig 4. Molecular assembly of (a)(a)(a)(a), (b) (b) (b) (b), and (c)(c)(c)(c) with POPC, a lipid representative of eucaryotic cell membranes. Calculated according to the Hypermatrix method2 . (a) (a) (a) (a) (b)(b)(b)(b) (c)(c)(c)(c) (a) (a) (a)
(a) (b)(b)(b)(b) (c)(c)(c)(c) Fig 5prediction of . In Silico the interactions of (a) (a)(a) (a), (b)(b)(b)(b) and (c) (c)(c) (c) with a model membrane calculated according to the IMPALA method3. Concentration (µmol/l) Concentration (µmol/l)Concentration (µmol/l) Concentration (µmol/l) (a)
(a) (a)
(a) (b)(b)(b)(b) (c)(c)(c)(c) Fig 6. Cell viability assay . Test based on the bioreduction of MTS into formazan (G5421, Promega) Fig 2. Adsorption isotherms recorded at 25°C
with a Langmuir film balance (KSV, Finland). Injection of pure compounds solutions into milliQ water (final concentration of about 6 µM)
1111
S TS TS TS T S T EPS T EPS T EPS T EP: S
: S
: S
: S
Y NTHESISY NTHESISY NTHESISY NTHESIS VIAVIAVIAVIA ANA NA N ENZYMATICA N ENZYMATICENZYMATICENZYMATIC R OUTER OUTER OUTER OUTEO C H3 O 12 O C H3 O OMe OH 12 (a) (a) (a)
(a) cinnamyl myristate
(c) (c) (c) (c) vanillyl myristate O C H3 O OMe OH 12 (b) (b) (b) (b) coniferyl myristate OH C H3 O 12 O H R 1 R2 n
+
Lipase CAL B 2-methyl-2-butanol, Molecular sieve, 60°C, 4h n=3 (+ 1 unsaturation) R1 = H R2 = H n=3 (+ 1 unsaturation) R1 = -OMe R2 = -OH n=1 (no unsaturation) R1 = -OMe R2 = -OH su bs tit ut io ns nu m be r o f c ar bo ns an d u ns atu ra tio n + H2O + H2O + H2OO
O
O
O
BJECTIVESBJECTIVESBJECTIVESBJECTIVESA structure-function relationship study was undertaken to identify the structural patterns affecting the interfacial and membrane interaction properties. A computational approach was used to predict the interactions
with a lipid and membrane model. The toxicity was evaluated via a cell
viability assay.
Specific antioxidant molecules (e.g. phenolic compounds) help to prevent oxidation reactions of the cell membrane. Grafting a long alkyl chain onto these compounds should improve their interfacial properties and their ability to interact with membrane lipids.
IIII
N TERESTSN TERESTSN TERESTSN TERESTSTable 1: Characteristic parameters of the monolayers determined by compression isotherms at air/water interface and by molecular modelling
A0 Ac ∏c Compressibility A,Modelling
(Ų/mole-cule) (Ų/mole-cule) (mN/m) (m/N) (Ų/mole-cule)
(a) (a)(a) (a) 48,6± 1,6 32,4± 0,9 8,2± 0,6 58,5 56,4± 1,7 (b) (b) (b) (b) 64,9± 1,0 23,2± 1,6 26,9± 4,0 82,1 (1) 27,6 (2) 57,3± 3,5 (c) (c) (c) (c) 70,9± 2,6 26,9± 1,3 31,4± 0,4 54,7 (1) 19,4 (2) 67,0± 3,7
Fig 3. Organization of the molecules at an hydrophobic/hydrophilic interface determinedin silico (Structure Tree method1).
(a) (a) (a) (a) (b)(b)(b)(b) (c)(c)(c)(c)
Toxicity evaluation
DiscussionThe addition of an alkyl chain confers interfacial properties to the phenolic compounds. (a)(a)(a),(a) (b)
(b) (b)
(b) and (c) (c) (c) (c) molecules have an affinity for the air/water interface; weak for (a)(a)(a) but (a) stronger and similar for (b)(b)(b)(b) and (c)(c)(c). They are able to form (c)
monolayers with increasing stabilities: (a)(a)(a)<(b)(a) (b)(b)<(c)(b) (c)(c)(c). We observe A0(a)(a)(a)(a)<A0(b)(b)(b)(b)<A0(c)(c)(c)(c):
molecular areas increase to maximize the amount of polar
groups in the hydrophile phase. When injected into the subphase, molecules absorbed
immediately onto the interface.
Discussion Molecule (a)(a)(a): non (a)
toxic. Molecules (b)(b)(b)(b) and (c)(c)(c): (c) toxicity dependant on concentrations and similar, Discussion For all molecules, the
ester bond neighbours POPC’s ester bond and the alkyl chains are close
and nearly parallel. (a)
(a)(a)
(a), (b)(b)(b) and (c)(b) (c)(c)(c) insert into the external
sheet of the membrane. The hydrophobic chains
are tilted. Polar phenolic groups of (b)
(b) (b)
(b) and (c)(c)(c)(c) remain in the hydrophobic
region and can probably cause membrane perturbations. Part (2)
Part (1)
References 1 Lins L., Biochemical Pharmacology, 52, 1155-1168, 1996. 2 Brasseur, R., CRC Press: Boca Raton, FL, Vol. 1, 203−219, 1990. 3Ducarme, Ph., Proteins: Structure, Function, and Genetics, 30, 357-371, 1998. @ 49 0n m Phi Pho Πc Αc Α0
