Cubic Phase Nanoparticles

Formulation and Characterization Formulation and Characterization of Cubosomes using Lipids and of Cubosomes using Lipids and

Block CopolymersBlock Copolymers

Ashish L. SarodeAshish L. SarodeMaster of ScienceMaster of Science

Biomedical and Pharmaceutical SciencesBiomedical and Pharmaceutical SciencesUniversity of Rhode IslandUniversity of Rhode Island

OutlineOutline

IntroductionIntroduction MaterialsMaterials MethodsMethods Results and DiscussionResults and Discussion ConclusionsConclusions

Dextromethorphan (DXM)Dextromethorphan (DXM)

ChemistryChemistry PharmacodynamicsPharmacodynamics PharmacokineticsPharmacokinetics Controlled ReleaseControlled Release

Dextromethorphan (DXM)Dextromethorphan (DXM)

Equivalence and DoseEquivalence and Dose 50 mg DXM Tannate = 30 mg DXM HBr50 mg DXM Tannate = 30 mg DXM HBr DXM Tannate Suspensions – 25 mg/5ml – 10 ml DXM Tannate Suspensions – 25 mg/5ml – 10 ml

every 6 hoursevery 6 hours Delsym – DXM Polistirex = 30 mg DXM HBrDelsym – DXM Polistirex = 30 mg DXM HBr Delsym – 10 ml every 12 hoursDelsym – 10 ml every 12 hours Controlled Release DXM Tannate Suspension – 50 Controlled Release DXM Tannate Suspension – 50

mg/5ml – 10 ml every 12 hoursmg/5ml – 10 ml every 12 hours

Cubic PhaseCubic Phase GeometryGeometry

Water channel diameter – 5 nmWater channel diameter – 5 nm Interfacial area – 400 mInterfacial area – 400 m22/g/g Thermodynamic stability – Thermodynamic stability –

curvature energy of each curvature energy of each monolayer vs. stretching energy of monolayer vs. stretching energy of amphiphile chainsamphiphile chains

Characterization – X-ray / neutron Characterization – X-ray / neutron diffraction, TEM, FT-IR, ESR, diffraction, TEM, FT-IR, ESR, NMR, DSC, and hot stage NMR, DSC, and hot stage microscopymicroscopy

Crystal lattices – double diamond Crystal lattices – double diamond (D-surface/Pn3m), body centered (D-surface/Pn3m), body centered (P-surface/Im3m), gyroid (G-(P-surface/Im3m), gyroid (G-surface/Ia3d)surface/Ia3d)

References: Shah et al., 2001; Wyatt and Dorschel, 1992

Reference: Spicer, 2001


Cubic PhaseCubic Phase LipidsLipids

Examples – monoolein or Examples – monoolein or glyceryl monooleate (GMO), glyceryl monooleate (GMO), monoelaidin, phospholipids, monoelaidin, phospholipids, PEGylated phospholipids, and PEGylated phospholipids, and phosphatidylethanolaminephosphatidylethanolamine

GMO phases – reversed GMO phases – reversed micellar (Lmicellar (L22), lamellar (L), lamellar (Lαα), ), reversed hexagonal (Hreversed hexagonal (Hππ), and ), and cubic (C)cubic (C)

Low water solubility (10Low water solubility (10-6-6M) of M) of GMO – cubic phase co-exist GMO – cubic phase co-exist in equilibrium with water (oral in equilibrium with water (oral controlled release)controlled release)

References: Shah et al., 2001; Larsson, 1989; Nielsen et al., 1998)

Cubic PhaseCubic Phase Block CopolymersBlock Copolymers

Tri-block copolymers Tri-block copolymers (-PEO-PPO-PEO-)(-PEO-PPO-PEO-)

Various phases depending Various phases depending on temperature and on temperature and concentrationconcentration

Pluronic F127 – lower Pluronic F127 – lower critical solution critical solution temperature below 37temperature below 37°C °C (20% solution)(20% solution)

Thermoreversible property Thermoreversible property – parenteral controlled – parenteral controlled releaserelease

Cubic phase can’t co-exist Cubic phase can’t co-exist in equilibrium with excess in equilibrium with excess water (no oral controlled water (no oral controlled release)release)

Reference: Hamley, 2004

CubosomesCubosomes

GeometryGeometry Dispersed particles of Dispersed particles of

cubic phasecubic phase Mathematical Mathematical

characterization by characterization by nodal surface nodal surface approximationsapproximations

Identification – Cryo-Identification – Cryo-TEM analysisTEM analysis


CubosomesCubosomes

ManufactureManufacture1.1. Bottom-up techniqueBottom-up technique

Formation and crystallization of cubosomes from Formation and crystallization of cubosomes from precursors on molecular scaleprecursors on molecular scale

2.2. Top-down techniqueTop-down technique Dispersion of cubic phases into cubosomes by high energy Dispersion of cubic phases into cubosomes by high energy

inputinput Energy sources – high pressure homogenization, and Energy sources – high pressure homogenization, and

sonicationsonication Formation of complex dispersions – vesicles and Formation of complex dispersions – vesicles and

cubosomescubosomes Aqueous solutions of block copolymers – low energy input, Aqueous solutions of block copolymers – low energy input,

prevent recoalescenceprevent recoalescence

ObjectiveObjective Recent discoveries in biotechnology and Recent discoveries in biotechnology and

genetics – proteins, peptides, plasmid DNA, and genetics – proteins, peptides, plasmid DNA, and oligonucleotidesoligonucleotides

Demand targeted delivery to specific cells – Demand targeted delivery to specific cells – need of drug delivery vehiclesneed of drug delivery vehicles

Lipidic cubic phase formulations is an optionLipidic cubic phase formulations is an option To acquire knowledge and experience of To acquire knowledge and experience of

working with lipids and polymers that are used to working with lipids and polymers that are used to make cubic phase formulationsmake cubic phase formulations

To investigate potential of cubosome delivery in To investigate potential of cubosome delivery in oral controlled releaseoral controlled release

MaterialsMaterials

InstrumentationInstrumentation High shear mixer and homogenizerHigh shear mixer and homogenizer USP Dissolution apparatus # 2USP Dissolution apparatus # 2 Phenomenex partisil column (10 SCX, Phenomenex partisil column (10 SCX,

4.6x250 mm, P/N00G-0127-E0)4.6x250 mm, P/N00G-0127-E0) HPLC-UV analytical systemHPLC-UV analytical system Cryo-TEMCryo-TEM

MaterialsMaterials

Chemicals and ReagentsChemicals and Reagents Glyceryl Monooleate (Myverol 18-92K, and Glyceryl Monooleate (Myverol 18-92K, and

18-99K) – Kerry Biosciences18-99K) – Kerry Biosciences Block Copolymers (Pluronic F-127, & L-64) – Block Copolymers (Pluronic F-127, & L-64) –

BASFBASF Oleic Acid – Fisher ScientificOleic Acid – Fisher Scientific Dextromethorphan Tannate, Dextromethorphan Tannate,

Dextromethorphan HBr, formulation Dextromethorphan HBr, formulation excipients, and analytical reagents – Hi-Tech excipients, and analytical reagents – Hi-Tech Pharmacal. Co., Inc.Pharmacal. Co., Inc.

Methods – FormulationMethods – FormulationIngredient

Formulation

F1 F2 F3 F4 F5 F6 F7

1. GMO 10 g 10 g 10 g 10 g 10 g 10 g 11 g

2. Oleic Acid 0 g 1 g 1 g 1 g 2 g 2 g 2 g

3. Dextromethorphan Tannate 0.5 g 0.5 g 0.5 g 1 g 1 g 1 g 1 g

4. Pluronic F-127 1 g 1 g 1 g 1 g 1 g 1 g 1.2 g

5. Purified Water 40 ml 40 ml 40 ml 40 ml 40 ml 40 ml 40 ml

6. Sucrose Syrup 40 ml 40 ml 40 ml 40 ml 40 ml 40 ml 40 ml

7. Propylene Glycol 2 g 2 g 2 g 2 g 2 g 2 g 2 g

8. Methylparaben 0.2 g 0.2 g 0.2 g 0.2 g 0.2 g 0.2 g 0.2 g

9. Propylparaben 0.02 g 0.02 g 0.02 g 0.02 g 0.02 g 0.02 g 0.02 g

10. Sucralose 0.02 g 0.02 g 0.02 g 0.02 g 0.02 g 0.02 g 0.02 g

11. Citric Acid 0.3 g 0.3 g 0.3 g 0.3 g 0.3 g 0.3 g 0.3 g

12. Sodium Citrate 1 g 1 g 1 g 1 g 1 g 1 g 1 g

13. Xanthan Gum 1 g 1 g 1 g 1 g 1 g 0.3 g 0.5 g

14. Sodium Alginate 0 g 0 g 0 g 0 g 0 g 0.15 g 0.1 g

15. Flavor qs qs qs qs qs qs qs

16. Color qs qs qs qs qs qs qs

17. Purified Water qs 100 ml qs 100 ml qs 100 ml qs 100 ml qs 100 ml qs 100 ml qs 100 ml

Note1: GMO for F3 is Myverol 18-99K, for all other formulations GMO is Myverol 18-92K

Note2: Actual formulations were prepared in 1000 ml

Note3: Concentration of the drug was then increased gradually in formulation F7 in the order 0.25% w/v, 0.5% w/v, 0.75% w/v, and 1% w/v

Methods – FormulationMethods – Formulation

MethodsMethods

In-vitro AnalysisIn-vitro Analysis Number of vessels – 6Number of vessels – 6 Medium – 0.1N HCl or 2%SDSMedium – 0.1N HCl or 2%SDS Volume – 900 mlVolume – 900 ml Temperature – 37Temperature – 37°C°C Speed – 50 RPM and 250 RPM for infinitySpeed – 50 RPM and 250 RPM for infinity

MethodsMethods

Quantitative AnalysisQuantitative Analysis Buffer – 0.087 M potassium phosphate Buffer – 0.087 M potassium phosphate

monobasic aqueous solutionmonobasic aqueous solution Mobile phase – methanol : buffer :: 4:6Mobile phase – methanol : buffer :: 4:6 Standard – dissolved in dissolution mediumStandard – dissolved in dissolution medium Injection volume – 10 Injection volume – 10 µlµl Flow Rate – 1.5 ml/minFlow Rate – 1.5 ml/min Run Time – 20 minutesRun Time – 20 minutes Wavelength – 205 nmWavelength – 205 nm

MethodsMethods

Cryo-TEM AnalysisCryo-TEM Analysis Final formulation (F7)Final formulation (F7) SamplesSamples

1.1. Overnight equilibrationOvernight equilibration2.2. Two weeks’ storageTwo weeks’ storage

Chamber humidity – 95%Chamber humidity – 95% Carbon grid – 100nm to 2Carbon grid – 100nm to 2µm pore diameterµm pore diameter Film thickness – 50 to 100 nmFilm thickness – 50 to 100 nm Equilibrated for 3 seconds – plunged in liquid Equilibrated for 3 seconds – plunged in liquid

ethane – stored in liquid nitrogenethane – stored in liquid nitrogen TEM images were taken at -170°CTEM images were taken at -170°C

Results and DiscussionResults and Discussion

In-vitro AnalysisIn-vitro Analysis

Dissolution Profiles (0.5% w/v of Dextromethorphan Tannate)

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F2 (in 0.1N HCl) F2 (in 2% SDS) F3 (in 0.1N HCl) F1 (in 0.1N HCl)


In-vitro AnalysisIn-vitro AnalysisDissolution Profile (1% w/v of Dextromethorphan Tannate)

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F4 (in 0.1 N HCl) F5 (in 0.1N HCl) F6 (in 0.1 N HCl) F7 (in 0.1 N HCl)



Dissolution Profiles

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0.25% w/v 0.50% w/v 0.75% w/v 1.0% w/v



Dissolution Profile (Delsym)

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Delsym (in 0.1 N HCl)



Dissolution Profile (Delsym vs. F7)

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Dissolution Profiles

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F7 Delsym


Cryo-TEM Analysis (overnight equilibration)Cryo-TEM Analysis (overnight equilibration)


Cryo-TEM Analysis (two weeks’ storage)Cryo-TEM Analysis (two weeks’ storage)

ConclusionsConclusions

Release of hydrophobic salts of some Release of hydrophobic salts of some drugs can be controlled by oral cubosome drugs can be controlled by oral cubosome deliverydelivery

Balance of cubosome structure, charge, Balance of cubosome structure, charge, and viscosity is important to achieve and viscosity is important to achieve controlled releasecontrolled release

Equivalent to suspensions of resin Equivalent to suspensions of resin complexed drugscomplexed drugs

AcknowledgementsAcknowledgements

Dr. Hossein ZiaDr. Hossein Zia Dr. Polireddy DondetiDr. Polireddy Dondeti Dr. Arijit BoseDr. Arijit Bose

ReferencesReferences Hamley, I. W. (2004). Introduction to Block Copolymers. In I. W. Hamley (Ed.), Developments in Hamley, I. W. (2004). Introduction to Block Copolymers. In I. W. Hamley (Ed.), Developments in

Block Copolymer Science and Technology: John Wiley & Sons, Ltd: Chichester.Block Copolymer Science and Technology: John Wiley & Sons, Ltd: Chichester. Larsson, K., 1989. Cubic lipid-water phases: structures and biomembrane aspects. J. Phys. Chem. Larsson, K., 1989. Cubic lipid-water phases: structures and biomembrane aspects. J. Phys. Chem.

93, 7301–7314.93, 7301–7314. Nielsen, S.L., Schubert, L., Hansen, J., 1998. Bioadhesive drug delivery systems. I. Characterization Nielsen, S.L., Schubert, L., Hansen, J., 1998. Bioadhesive drug delivery systems. I. Characterization

of mucoadhesive properties of system based on glyceryl mono-oleate and of mucoadhesive properties of system based on glyceryl mono-oleate and glyceryl mono-linoleate. glyceryl mono-linoleate. Eur. J. Pharm. Sci. 6, 231–239.Eur. J. Pharm. Sci. 6, 231–239.

Shah, J.C., Sadhale, Y., Chilukuri, D.M., 2001. Cubic phase gels as drug delivery system. Adv. Shah, J.C., Sadhale, Y., Chilukuri, D.M., 2001. Cubic phase gels as drug delivery system. Adv. Drug Deliv. Rev. 47, 229–250.Drug Deliv. Rev. 47, 229–250.

Spicer, P. T., Hayden, K. L., Lynch, M. L., Ofori-Boateng, A., Burns, J. L. (2001). Novel Process Spicer, P. T., Hayden, K. L., Lynch, M. L., Ofori-Boateng, A., Burns, J. L. (2001). Novel Process for Producing Cubic Liquid Crystalline Nanoparticles (Cubosomes). Langmuir, 17, 5748-5756.for Producing Cubic Liquid Crystalline Nanoparticles (Cubosomes). Langmuir, 17, 5748-5756.

Spicer, P. T. (2004). Cubosomes: Bicontinuous Cubic Liquid Crystalline Nanostructured Particles, Spicer, P. T. (2004). Cubosomes: Bicontinuous Cubic Liquid Crystalline Nanostructured Particles, Dekker Encyclopedia of Nanoscience and Nanotechnology (Vol. 1, pp. 881-892): Marcel Dekker.Dekker Encyclopedia of Nanoscience and Nanotechnology (Vol. 1, pp. 881-892): Marcel Dekker.

Spicer, P. T. (2005). Progress in liquid crystalline dispersions: Cobosomes. Current Opinion in Spicer, P. T. (2005). Progress in liquid crystalline dispersions: Cobosomes. Current Opinion in Colloid & Interface Science, 10, 274-279.Colloid & Interface Science, 10, 274-279.

Wyatt, D., Dorschel, D., 1992. A cubic-phase delivery system composed of glyceryl monooleate and Wyatt, D., Dorschel, D., 1992. A cubic-phase delivery system composed of glyceryl monooleate and water for sustained release of water-soluble drugs. Pharm. Technol. 16, 116–130.water for sustained release of water-soluble drugs. Pharm. Technol. 16, 116–130.

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Cubic Phase Nanoparticles