Preparation and

Characterization of Copolymer
Micelles for the Solubilization
and In Vitro Release of Luteolin
and Luteoloside
Pharmacutical nanotechnology
Masters of industrial pharmaceutical technology
Prepared by : Yara Al-Salhi
Submitted to : Dr. Moammal Qurt
Contents

 Issue
 Drug
 Diblock copolymer micelles
 Copolymers
 Characterization of copolymers
 Preparation of drug-Loaded Copolymer Micelles
 Results and discussion
 Conclusion
Issue of consideration

 Luteolin and luteoloside belong to flavonoids, both have high potentials in

diverse treatments.
 However, their clinical applications are largely hampered because of poor
solubility, low bioavailability, and oral absorption.
 Encapsulation of hydrophobic drugs into block copolymer micelles is a
promising approach to improve the drug solubility and bioavailability.
Drug

 Luteolin is a flavonoid.
 It has anticancer, anti-inflammatory, anti-allergic, and anti-amnesic activities.
 Luteoloside is the glucoside of LUT with a 7- O-β-glucose moiety in the
molecular structure
 LUS has anti-inflammatory, anticancer, anti-ischemic, and anti-HBV effects.
Diblock copolymer micelles
 Diblock copolymer micelles have a hydrophobic core as a drug carrier and a
hydrophilic shell as an invisible cloak to the body defense system, which can
prolong circulation time in vivo
Copolymers

 Methoxy polyethylene glycol-polycaprolactone (mPEG5K-PCL10K)

 Methoxy polyethylene glycol-polylactide-co-glycolide (mPEG5K-PLGA10K)
 Methoxy polyethylene glycol-polylactide (mPEG5K-PDLLA10K)
 mPEG-PCL, mPEG-PLGA, mPEG-PDLLA : are all diblock copolymers, which are
biodegradable and biocompatible
Characterization of Copolymers

 The Molecular Weight of Copolymers

 CMC and Stability of Copolymers
 The Micropolarity of LUT (or LUS) in Copolymers Micelles
 Experimental Design and Optimization by RSM
Preparation of LUT (or LUS)-Loaded
Copolymer Micelles
 The diblock copolymers are used to load the drug by a self-assembled method.
 Briefly, the drug and mPEG5K-PCL10K copolymer mixture were co-dissolved in 6 mL of THF.
 Next, the solution was dropwise dispersed in 10 mL of water under ultrasound at 50 Hz for 40
min.
 The solution was continuously stirred for 48 h in order to ensure that THF was evaporated.
 In this process, mPEG5K-PCL10K self-assembled into core/shell structured micelle with core
encapsulated LUT (or LUS).
 The LUT- (or LUS)-loaded mPEG5K-PCL10K micelle solution was filtered through a 0.45- μm
membrane.
 Eventually, a clear and homogeneous yellow drug micelle solution was obtained and stored
at 4°C.
 LUT- (or LUS)-loaded mPEG5K-PLGA10K (or mPEG5K-PDLLA10K) micelle solution was similar
to the steps above, except for acetone instead of THF.
The Solubility of LUT (or LUS) in Water
and Copolymer Micelles
 The solubility were determined by UV-Vis spectrophotometer according to the
method of Yan with modification.
 Briefly, 10 mL of distilled water and excessive LUT (or LUS) were placed in a 50-mL
conical flask and shaken and balanced to make drug fully dissolved for 24 h at 37°C.
 Next, the solution was centrifuged at high speed for 10 min, and supernatant was
filtered through a 0.45- μm membrane.
 The filtrate was diluted with water to 25 mL.
 The absorbance of the drug was analyzed at 354 nm, and solubility in water was
calculated.
 Similarly, the solubility of the drug in copolymer micelles was also calculated.
 In order to obtain the representative data, the results were recorded as the
averages plus or minus standard deviation (n = 3, ±SD).
Characterization of Drug Micelles

 The concentration of LUT (or LUS) loaded into diblock copolymer micelles
were determined by diluting the micellar dispersions with EtOH and the
absorbance measured at 354 nm on a UV-Vis spectrophotometer.
 The entrapment efficiency (EE) and loading capacity (LC) were calculated.
 The average size and zeta potential of drug micelles were obtained from
three repeat measurements on Malvern NanoZS90 instruments.
 The results were recorded as the averages plus or minus standard deviation (n
= 3, ±SD).
 During the measuring process, the temperature was kept at 25°C.
 Drug micelles were placed on carbon-coated copper grids (200 mesh), and
morphologies were observed.
In Vitro Drug Release Study

 In order to determine the release profile of the drug ,the freeze-dried powder
of drug micelles equivalent of 0.15 mg of drug was dissolved in 10 mL
phosphate-buffered solution (PBS) and placed in a dialysis bag.
 The dialysis bag was incubated in 80 mL of PBS at 37°C with gentle shaking.
At predetermined time points, 3 mL of release medium was withdrawn and
the same volume of fresh PBS solution was added to maintain the constant
volume.
 The amount of released was determined at a wavelength of 354 nm using the
UV-Vis method.
 The accumulative percentage release (Q%) was calculated
Results and discussion
The Molecular Weight of Copolymers
 The samples were dissolved in chromatographic (THF) at room temperature
for 6 h with 0.45-μm filtration. The chromatographic column is MZ-Gel SDplus,
mobile phase is THF, and the flow rate is 1.000 mL min−1
 The weights (Mw) of mPEG5K-PCL10K = 21,700
 mPEG5K-PLGA10K = 19,040
 and mPEG5K-PDLLA10K = 12,060
 And dispersion indexes (Mw/Mn) are 1.524, 1.583, and 1.524, respectively,
which is in the ideal range
CMC, Stability of Copolymers and Micropolarity
The Optimization of Drug Micelle
Formulation
 THF, acetone, CHCl3, and CH2Cl2 are selected to prepare the blank micelles,
because the average size of nanomicelles is affected by organic solvents.
 All of diblock copolymers can form opalescence transparent solution in four
organic solvents under ultrasound at 50 Hz.
 Average size of mPEG5K-PCL10K blank micelle in THF is 67.8 nm which is
smaller than other three solvents with better PDI.
 However, the sizes of both mPEG5KPLGA10K and mPEG5K-PDLLA10K blank
micelles in acetone are 57.3 and 66.8 nm, respectively, which are all smaller
than the other three solvents with better PDI.
The Solubility of LUT and LUS
Characterization of Drug Micelles
In Vitro Drug Release Study
CONCLUSION

 The diblock copolymers have a certain solubilization effect on LUT and LUS.
 LUT- (or LUS)-loaded mPEG5K-PCL10K exhibited a better stability and encapsulation
efficiency, and mPEG5K-PLGA10K exhibited higher loading capacity for LUS.
 The obtained drug micelles were torispherical with average diameter about 70 nm.
 The in vitro release behavior of LUT (or LUS) from the micelles showed the preparation
possessed a sustained release property.
 The accumulative percentage release of LUT (or LUS) from the micelles in pH 7.4 PBS
solution was higher than that in pH 6.4 PBS solution in 48 h.
 mPEG5K-PCL10K and mPEG5K-PLGA10K can become a potential nanocarrier for LUT and
LUS to overcome their poor solubility in water. Besides, the obtained LUT (or LUS) micelles
could be lyophilized into a powder form without any other excipient and cryoprotector,
while the re-dissolved LUT (or LUS) micelles are homogeneous and stable. So they are
expected to be made into oral or intravenous formulations for clinical application.
 The end
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