Towards Better Photosensitizers: Synthesis and Characterization of New

Tetraphenylporphyrins

By: Deborah Zhao

Mentors: Dr. Ursula Simonis

Sherry Farokhzad

What is a Porphyrin?

A porphyrin is a heterocyclic (a substance that contains a ring structure) macrocycle (any molecule that contains a ring of large numbers of atoms) that’s made from three pyrrole subunits and one pyrroline subunit which are linked on by the opposite sides through four methine bridges.

Tetraphenylporphyrin Porphyrin

http://omlc.ogi.edu/spectra/PhotochemCAD/str_gif/TPP http://www.answers.com/main/content/wp/en/thumb/f/f7/250px-Porphyrin-structure.png

Introduction to Photodynamic Therapy (PDT)

• PDT is a treatment that uses the application of a photosensitizing agent, which is then illuminated by a light source with presence of molecular oxygen to destroy unwanted tissue.

• A photosensitizer is a substance that absorbs energy from light and uses that energy to enable chemical reactions to happen (i.e. to destroy unwanted tissue).

• Porphyrin-based compounds are most widely studied in the use of photosensitizers because prophyrins can strongly absorb visible light.

• PDT is used to treat many serious disease such as: cancers, skin lesions, age-related macular degeneration, and coronary artery disease.

How Does PDT Work With Porphyrins?

1. A porphyrin photosensitizer is injected into the patient (it can also be applied topically if treating external area).

2. After time, the photosensitizer will accumulate into the unwanted tissue.

3. Careful illumination of a laser light with an approximate wavelength of 750 nm will cause reactive oxygen species (ROS) to form which damages the cell’s organelles therefore causing apoptosis (cell death).

http://cal.it.nuigalway.ie/incl/medical6.gif

What are the Advantages and Disadvantages of PDT?

• Essentially painless, therefore it’s well tolerated.• It’s easily performed by specialists as compared to the

complexity of surgery.• The treatments are less invasive than surgery.• Scarring risks are reduced and cosmetic outcomes are

improved.• The photosensitizers accumulates more into the

abnormal tissue but there is still a small amount in the normal tissue.

• The side effects are minimal but patients cannot be in direct sunlight.

• Sometimes prolonged photosensitivity and phototoxicity can occur.

• Still in need of a better photosensitizer.

The Ideal Photosensitizer

• It has to be pure so it doesn’t have any side reactions.• Easily synthesized and composed.• It can produce a high quantum yield to reactice oxygen

species.• It has an absorbance anywhere between 650-1000 nm

so it can penetrate into deep tissues.• It’s water-soluble so it can easily travel through the

bloodstream.• It has rapid body clearance to avoid prolonged

photosensitivity.

Currently Drug-Approved Photosensitizers

N HN

NH N

NaO2CNaO2C

N HNNH N

HO2CH3CO2C

H3C

H3CO2C

H3CO2CO

O

n = 1-9

HO NH2

O

O

N HN

NH N

HOOCHOOC

Photofrin Visudyne

Protoporphyrin IX

Levulan or ALA

N HN

NH N

Foscan

OH

OH

OH

HO

• None of these drugs fulfills the job of an ideal photosensitizer.

• Some of the drugs are unable to absorb much red light therefore unable to penetrate into deep tissues.

• Some of the drugs have slow clearance therefore causing prolonged photosensitivity or phototoxicity.

The Goal of Our Project

Find out if tertraphenylporphyrins with the addition of serine will become a better photosensitizer by:

• Purifying and reducing a mixture of tetraphenylporphrins.

• Extract the cis (p-F)2(p-NH2)2TPPH2 and add on the serine.

• Characterize the prophyrin with 1H NMR spectroscopy.

Why cis (p-F)2(p-NO2)2TPPH2?

Synthesizing the Tetraphenylporphyrins

N HNNH N

F

NO2

F

O2N

N HNNH N

NO2

F

F

O2N

F

H

O

NO2

H

O

NH

1 1 2+ +

4-Fluorobenzaldehyde 4-Nitrobenzaldehyde Pyrrole

cis(p-F)2-(p-NO2)2TPPH2 trans(p-F)2-(p-NO2)2TPPH2

+

+ 4 other compounds

Reflux at 80 oC

Proprionic acidChloroform

Adler et al., J. Org. Chem., 1967, 32, 476

Purifying and Separating the Porphyrins

• Column chromatography was used to purify the mixture of porphyrins by removing the tar and separating the different bands of porphyrin compounds.

• Loading the column and separation of the bands:

• Silica gel was used for the stationary phase and toluene was used for the mobile phase.

• Since silica is a polar compound, the less polar porphyrins will elute first while the more polar porphyrins will elute last.

1. TetranitroTPP

2. Trinitro-mono-fluoroTPP

3. & 4. Cis and Trans-difluoro-dinitroTPP

5. Trifluoro-mono-nitroTPP

6. TetrafluoroTPP

Tin Chloride Reduction of Nitro groups to Amines

N HNNH N

F

NO2

F

O2N

N HNNH N

NO2

F

F

O2N

SnCl2 x 2H2O

12 M HCl, 15 hours

N HNNH N

F

NH2

F

H2N

N HNNH N

NH2

F

F

H2N

Collman et al., J. ACS, 1980, 12, 102

Monitoring the Purification Process

Thin Layer Chromatography-TLC• A TLC plate is similar to a column, it compares the compounds by its polarity.

• Since the TLC plate is made of silica, the less polar compounds will travel closer to the solvent front.

• TLC is a comparative test, it does not show any information regarding the compounds’ molecular structure.

• TLC can also be used to test a compound’s purity.

http://sms.kaist.ac.kr/~jhkwak/gc/catofp/chromato/tlc/pic/004634st.jpg

Addition of Protected Serine

• Based on the known properties of cis and trans (p-F)2(p-NH2)2TPPH2, the addition of serine can possibly make the porphyrins somewhat water-soluble therefore making it a better photosensitizer because it can easily travel through the bloodstream.

Characterizing the Porphyrins

Results and Discussion

Future Works

Acknowledgements