MULTICOMPONENT COMPLEXATION OF DEHYDROEPIANDROSTERONE
WITH -CYCLODEXTRIN AND GLYCINE
M.Cirri1, P. Mura1, P. Corvi Mora2, F. Carli3
1
Dip. Scienze Farmaceutiche, Univ. Firenze, Via Capponi 9, 50121 Firenze, Italy
2
EUPHAR GROUP srl, Via Gandine 4/6, 29100 Piacenza, Italy
3
ACTIMEX srl, via Flavia 23/1, 34138 Trieste, Italy
Dehydroepiandrosterone (DHEA) is an adrenal steroid hormone that naturally decrease with
aging. DHEA replacement therapy in elderly people appears to have remedial effects against several
diseases of aging, enhancing quality of life with no significant adverse effects [1]. Moreover,
DHEA treatment showed antidepressant and promemory effects [2] and improved well-being and
sexuality in women with adrenal insufficiency [3]. However, the very limited aqueous solubility of
DHEA can give rise to both problems of formulation and low and variable bioavailability.
Cyclodextrin complexation has proved to be an effective method for enhancing dissolution
properties and bioavailability of poorly water-soluble drugs. Furthermore, recent studies reported
that the addition of a suitable third component can often significantly improve the cyclodextrin
solubilizing efficiency towards drugs.
Therefore, the aim of this work was to improve the DHEA bioavailability by
multicomponent complexation. Preliminary solubility studies enabled to select -cyclodextrin and
glycine as the most effective components for the ternary complex [4]. Solid DHEA/cyclodextrin/glycine systems in different molar ratios were prepared by mechanochemical
activation, by cogrinding physical mixtures in a high energy mill, and characterized by DSC, IR and
X-Ray powder diffraction analyses. The 1:2:3 (mol/mol) drug:Cd:glycine ternary system gave the
best results in terms of solubility and dissolution properties (USP paddle method) and was then
selected for in vivo studies. A first series of experiments were performed on 6 volunteers healthy
aged (83.3±4.6) men. The kinetic of native DHEA was compared with that of DHEA as ternary
complex (C-DHEA) after an oral load of 25 mg drug, orally randomly administered with a 4 days
interval. Administration of the drug as multi-component system resulted in enhanced bioavailability of DHEA, without affecting sulpho-conjugation nor bioconversion to testosterone [5].
Further in vivo studies were then carried out to confirm these preliminary results. Sixteen
women, suffering by adrenocortical insufficiency, aged between 18-75 years, participated in this
study. The DHEA kinetics was monitored after treating the volunteers with an oral load of 25 mg
of native DHEA or an equivalent amount of tc-DHEA formulated in tablets. The two treatments
were randomized and crossed over. A washout of four days separed each treatment. The patterns of
serum DHEA and DHEA sulphate levels after tc-DHEA administration were significantly higher,
both as Cmax (P <0.001) and AUC (P <0.05) values, than after untreated DHEA administration.
Moreover, the mean total time having DHEA serum levels within physiological range (> 3.9
ng/ml) was more than tripled using drug as ternary complex compared to the same dosage of native
DHEA. No significant difference was instead observed in DHEA sulphate/DHEA molar ratio
throughout the study period by comparing the values obtained after DHEA or tc-DHEA
administration, indicating that this latter proportionately enhanced the serum circulating levels of
both DHEA and DHEA sulphate (Data to be submitted for publication).
Finally, in order to shed light on the favourable role of glycine in improving -Cd
solubilizing efficiency towards DHEA, molecular modelling studies were performed. Analysis and
modelling of the structures of single components, binary and multicomponent complexes were
carried out using the INSIGHT 2.2.0 program from Biosym Technologies. The molecular structure
of -Cd was obtained from crystallographic parameters provided by the Cambridge
Crystallographic Data Centre. Conformational energy was evaluated applying the Discover
program, version 97.0 (Biosym/MSI). The force field calculations were performed using the
AMBER method suitably modified by adding specific parameters for carbohydrates according to
Homans [6].
The stability of the supramolecular systems was correlated to the value of the docking energy
between the interacting molecules that can be considered as a sum of van der Waals’ and
electrostatic energies among all the complex atoms. It was calculated as the difference between the
total complex energy and those of its components and it represents the gain of potential energy due
to interactions between the molecules forming the complex. The comparison between the docking
energy values obtained for the most stable conformations of DHEA--Cd complex (-14 kcal/mol)
and 1:1:2 DHEA--Cd-glycine complex (-44 kcal/mol) confirmed the stabilizing role of glycine
molecules on DHEA--Cd inclusion complexation. The results of molecular modelling studies
suggested the possible interactions, through hydrogen bonding, of the carboxylic groups of two
glycine molecules with two secondary hydroxyl groups on the wider side of cyclodextrin rim, thus
partially surrounding the part of the steroid not included in the host cavity and thus bettering its
wettability and dissolution properties. Moreover molecular modelling calculations also indicated the
possible formation in solution of more complex structures (1:2:3 DHEA--Cd-glycine) where a
DHEA molecule was interposed between two -Cds, with its external rings located at the entrance
of the wider rims of Cd molecules; the structure is further stabilized by glycine molecules which
place themselves as go-betweens for -Cd molecules, interacting throught hydrogen bonds between
their carboxylic groups and the Cd hydroxyl groups [7].
In conclusion, molecular modelling studies accounted for the important role of glycine in
ternary DHEA complexation with -Cd. However, further studies (through NMR spectroscopy and
synchrotron-radiation potentiated techniques such as SAXS, SuperESCA and X-ray powder
diffractometry) will be necessary to search for a confirmation to these preliminary findings and
obtain experimental data to utilize for elaboration of geometric models of the multicomponent
complex through molecular modelling, thus enhancing the model reliability.
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