pH-Sensitive Liposomes for Effective Release of Therapeutics

By: Savita RamlallMentor: Professor Sofou & Shrirang Karve

Polytechnic University

Laboratory of Drug Delivery Systems

What are Liposomes?A liposomes is a spherical vesicle with a membrane composed of a phospholipid bi-layer used to deliver drugs or genetic material into a cell.

When using liposomes as drug carriers the main aim is to reduce toxic side affects in sensitive organs such as the heart and brain and to target specific tissues such as cancer cells.

The Purpose of This Project

It involves lipid separation which leads to imperfections in membrane. This leads to leakage of drugs.

Materials

Prepared liposomes

Pipettes

Fluorescence Spectrometer

ProceduresTake Lipids in (CHCl3 solution) in a round

bottom flask dried under N2 for 5 minutesAnneal it with PBS solution for 2 hoursExtrude it through 100 nm membraneMeasure the intensities for excimer and

monomer for pH 7.4; 5.5; 4.0

Leakage mechanism: lipids with pyrene-labeled fatty acid chains to detect membrane asymmetry and destabilization

ex: 344 nm/ em: 396 nm ex: 344 nm/ em: 470 nm

time (minutes)10 100 1000

% c

onte

nt r

eten

tion

-60

-40

-20

0

20

40

60

80

100

pH = 7.4 pH = 5.5pH = 4.0

day 2day 3

Release of contents from rigid PEGylated liposomes vs pH (in PBS)

• pH dependent content release

• size of ‘lipid-packing’ discontinuities (3kDa dextrans are released similarly, 10kDa dextrans are retained at all pH values)

Release of contents from rigid PEGylated liposomes vs pH (in media with 10% serum)

time (minutes)10 100 1000

% c

ont

ent

ret

ent

ion

-60

-40

-20

0

20

40

60

80

100

day 2 day 3

pH = 7.4 pH = 5.5pH = 4.0

• Instantaneous content release at pH 4• significant release at pH 5.5 • stable content retention at pH 7.4 the first hours

Monomer-to-eximer ratio

day 1 day 2 day 3

pH = 7.4 0.64 ± 0.05 0.56 ± 0.08 0.57 ± 0.05

pH = 5.5 0.62 ± 0.06 0.52 ± 0.08 0.45 ± 0.06

pH = 4.0 0.58 ± 0.05 0.40 ± 0.05 0.42 ± 0.11

monomer-to-eximer ratio raft formation

Result & DiscussionContent release at different pH in PBS & media (10% serum) studied using calcein leakage.Higher content release at lower pHMembrane asymmetry at different pH was studied using pyrene monomer – excimer ratio.Asymmetry increases with lower values of pH.

Further StudiesDuring the remaining summer I would like to see the effect of temperature on the raft formation of the liposomal membranes and compare it with the effect of pH levels.

References

http://biophysics.org/discussions/2004/london.pdf

http://www.ajhp.org/cgi/content/abstract/46/8/1576

http://explantion-guide.info/meaning/liposome.html

Ulrich, A Biophysical Aspects of Using Liposomes as Delivery Vehicles vol. 22, NO. 2 (2002) 129-146

Torchilin, V Fluorescence microscopy to follow the targeting of liposomes and micelles to cells and their intracellular fate. (2004) 96-107

Lasic, D Noel applications of liposomes vol. 16 (1998) 307-319

AcknowledgementsProfessor Sofou

Shrirang Karve

Polytechnic University

Everyone in the Laboratory of Drug Delivery Systems

Dr. Sat

Harlem Children Society

The Audience

Special Thanks

LAB