order to avoid the methodological problems that were observed with the
conventional release methods, transfer experiments from the donor monoolein
dispersions to lipophilic acceptor MLV were used instead of these conventional
release techniques. The lipophilic acceptor MLV are intended to mimic
lipophilic compounds present in the blood, e.g. lipoproteins or cellular
structures and thereby present a compartment in which the released lipophilic
substance is soluble.
detection of the substances transferred from the monoolein dispersions into the
acceptor MLV was performed using two techniques. In case of the centrifugation technique one of the two
populations (donor or acceptor) should be in the liquid state while the other
should be in the crystalline form to be easily separated from each other. The
acceptor MLV was prepared with a highly concentrated sucrose solution in order
to be easily separated as pellet after centrifugation. The use of egg phosphatidyl choline and
cholesterol in the preparation of the acceptor MLV was to resemble many
physiological membranes. As an advantage with the flow cytometric technique,
it does not require a separation step
between the donor and acceptor particles and thus it showed an easier
methodological process than the centrifugation technique. Although this advantage of the flow
cytometric technique it requires the use of fluorescent substances as drug
model and large acceptor particles to be detected by a flow cytometer. On the other
hand, the donor particles should have a small particle size (less than 0.5 µm) in order to avoid the interference
with the acceptor MLV during the measurements.
The differences in the
transfer course (rate and amount) between the donor monoolein vesicles and
monoolein cubic particles could be attributed to the sponge like structure of
the monoolein cubic particles which decreases the rate and amount of drug
transferred from this structure. These results are in agreement with previous
observations (24), which indicate that cubic
particles should be quite useful for a rapid uptake because they can rapidly absorb pollutants (e.g., for water treatment or cosmetic skin
protection) and retain an amount determined by the solute partition
These observations supported the high affinity of the lipophilic porphyrin to
the donor monoolein cubic particles and consequently the lower amount and rate
of transfer that were observed from such donor particles. Further explanation
to this high affinity to the donor cubic particles is the unique structure of
these donor particles with a high specific bilayer/water interfacial area
(500-600 m2/g lipid) (4). As mentioned before (25), the interfacial area
(lipid/water interface) plays an important role in the transfer of porphyrin to
the different acceptor particles. This high bilayer/water interfacial area of
the cubic particles decreases the diffusion of the drug molecules outside the
donor cubic particles and subsequently a slow rate of drug transfer was
it was reported that the transfer of temoporfin, which is a porphyrin structure
drug molecule, was limited only to the interface of the acceptor liposomes
(were not entrapped in the vesicles bilayer) (16, 25)
which means that after saturation of this interface the transfer stopped at low
values. These recent findings might explain the low equilibrium values, which
were observed in the transfer of porphyrin from both donor particles (monoolein
vesicles and cubic particles), to the acceptor MLV. Furthermore, the acceptor
MLV had a large particle size which will be saturated with small amount and
thus probably the transfer stopped at a low level. Additionally, the acceptor
MLV were prepared from EPC with the addition of cholesterol, which increases
the rigidity of the bilayer (26) and occupies a part of the accessible outer surface and so
decreased the amount of drug transfer to the acceptor liposomes. Increasing the
acceptor to donor ratio from 1:25 to 1:100 led to an increase in the final
percent of drug transferred and this may be attributed to the increase in the
number of the acceptor particles relative to the donor particles, which in turn
increases the accessible surface available for drug transfer.
Finally both techniques nearly showed the same
results of transfer behaviour with a slight exception of the transfer rate
which was lower with the flow cytometric technique in comparison with the
centrifugation technique. This slight difference in the transfer rate constant
could be attributed to the separation step which was essential with the
centrifugation technique. This separation took time and during this time the
transfer can continue which led to an overestimation of the transfer rate
constant. This overestimation was
overcome with the flow cytometric technique. Also the flow cytometric
methodology was easier than the centrifugation due to the absence of this
to the conventional release methods, the transfer to a lipophilic acceptor
compartment as multilamellar vesicles is better than the commonly applied
release methods relative to the conditions in the blood. Both techniques are
suitable techniques to study the drug release from monoolein dispersions. Additionally
these techniques avoid the methodological problems which were observed with the
conventional release methods. Monoolein dispersions containing cubic
particles or vesicles can very successfully be used as a drug carrier.
Declaration of no conflict of interest
There are no conflicts of interest.
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