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SYNTHESIS, SPECTRAL CHARACTERIZATION AND BIOACTIVITY OF NOVEL ORGANOPHOSPHORUS
COMPOUNDS
DEPARTMENT OF BIOCHEMISTRY AND MICROBIOLOGY
CANDIDATE: NEKHUMBE DIIMISENI EZEKIEL
SUPERVISOR: Prof. AR Opoku
Co-supervisor: Dr. C Sampath
INTRODUCTION TO ORGANOPHOSPHORUS COMPOUNDS
Organophosphorus compounds are chemical compounds containing carbon-phosphorus bonds.
Phosphorus-carbon bond formation has attracted much attention because of its application in organic synthesis and bioorganic chemistry.
Organophosphorus compounds are mostly esters, amides or thiol derivatives of phosphonic acid and constitute a large group of chemical compounds.
APPLICATIONS
VERTERINARY MEDICININE PHARMACEUTICALS
AGRICULTURE - INSECTICIDES
INTRODUCTION TO ALPHA-HYDROXYPHOSPHONATES
What are α-Hydroxyphosphonates?
α- Hydroxyphosphonates are class of Organophosphorus compounds of synthetic interest because of their biological activity.
R
HO
P
OR1
OR1
O
ALPHA CARBON
Their biological activities include: anti-cancer, anti-viral, anti-bacterial, anti-fungal, anti-leukemic and many other biological activities (Subba et al., 2012).
They are useful precursors of α- functionalized phosphonates such as:-
α-Aminophosphonates
α-Aminophosphonic acids Pudovik reaction(Pudovik, 1972)
Kabachnik-Field reaction(Fields, 1952)
The two main routes for synthesis of α-Hydroxyphosphonates Pudovik reaction Abramov reaction
In the Abramov reaction, an aldehyde or a ketone is heated with trialkyl or dialkylphosphite to obtain alpha-Hydroxyphosphonates (Subba et al., 2012).
AIM OF THE CURRENT RESEARCH PROJECT
To synthesize Novel α-Hydroxyphosphonates compounds and test their antibacterial and anticancer activities.
OBJECTIVES OF THE PROJECT
Design and synthesize eco-friendly novel α-Hydroxyphosphonates having biological importance.
Develop simple and effective method for synthesis of target molecules.
Characterize the structure of the newly synthesized α-Hydroxyphosphonates by elemental analysis, spectral techniques.
Assess the antibacterial and anticancer activity of the synthesized compounds.
MATERIALS AND METHODS
RCHO + P
O
OR1H
OR1P
O
OR1
OR1
OH
R
Dialkylphosphite Alpha Hydroxyphosphonates
Here R= Various aldehyedes R1= Phenyl, ethyl, methyl, iso propyl, butyl, etc
Reflux for 4-5 hours
1,4-Dimethyl piperizine
Synthesis of Alpha-Hydroxyphosphonates compounds by Abramov reaction.
RESULTS AND DISCUSSION
.COMPOUND 1
O2N CHO + PHO2N C P O
1,4-Dimethyl piperizine
THF (680C), Ref lux 4-5 hours
OH
H
O
4- Nitrobenzaldehyde Diphenyl phosphite
TLC system:- 7:3 Hexane and ethyl acetateYield:- 80%
O
O
Diphenyl (hydroxy(4-nitrocyclohexyl)methyl)phosphonate
O O
MP: 80-82oC
CHO
F
COMPOUND 2
+ PH
O
F
C
OH
H
P O
3-Flourobenzaldehyde Diphenyl phosphite
1,4- Dimethyl piperizine
THF (680C), Ref lux 4-5 hours
TLC system:- 7:3 Hexane and ethyl acetateYield:- 72%
O
OOO
diphenyl ((3-f luorocyclohexyl)(hydroxy)methyl)phosphonate
MP: 74-76oC
COMPOUND 3
NH
CHO
+ PH
O
NH
C HHO
P
O
1,4-Dimethly piperizine
THF (680C), Ref lux 4-5 hours
4-pyridinebenzaldehyde Diphenyl phosphite
TLC system:- 7:3 Ethly acetate and hexane (suitable system) 7:3 Hexane and ethyl acetate (not suitable system)Yield:- 78%
O
O O
O
(R)-diphenyl (hydroxy(piperidin-4-yl)methyl)phosphonate
MP: 130-132oC
NH
+ HP
O
NH
C
OH
H
P
3-Pyridine aldehyde Diphenyl phosphie
COMPOUND 4
1,4-Dimethyl piperizine
THF (680C), Reflux 4-5 hours
TLC system:- 7:3 Hexane and ethyl acetate
OCHO
O O
O
O
Diphenyl (hydroxy(piperidin-3-yl)methyl)phosphonate
MP: 142-144oC
COMPOUND 5
Cl
CHO
+ PH
O
Cl
C HHO
P
O
1,4-Dimethyl piperizine
THF (680C), Reflux 4-5 hours
4-ChlorobenzaldehydeDiphenyl phosphite
TLC system:- 7:3 Hexane and ethyl acetateYield:- 71%
O O
OO
(R)-diphenyl ((4-chlorocyclohexyl)(hydroxy)methyl)phosphonate
MP: 60-69oC
COMPOUND 6
Cl
Cl CHO + HP
O
C P O
OH
H
Cl
Cl
1,4-Dimethyl piperizine
THF(680C), Reflux 4-5 hours
Diphenyl phosphite2,4-Dichlorobenzaldehyde
TLC sytem:- 7:3 Hexane and ethyl acetateYield:- 70%
O O
O
O
diphenyl ((2,4-dichlorocyclohexyl)(hydroxy)methyl)phosphonate
MP: 73-75oC
COMPOUND 7
O
O
CHO
+ PH
O O
O
C
H
OH
P O
1,4-Dimethyl piperizine
THF (680C), Ref lux 4-5 hours
Piperonaldehyde Diphenyl phosphite
TLC System:- 7:3 Hexane and ethly acetateYield:- 73%
O
O
OO
diphenyl ((hexahydrobenzo[d][1,3]dioxol-5-yl)(hydroxy)methyl)phosphonate
COMPOUND 8
NH
CHO
+ PH
O
NH
C
H
OH
P O
1,4-Dimethly piperizine
THF (680C), Reflux 4-5 hours
Diphenyl phosphite
Indole-3-aldehyde
TLC System:- 7:3 Hexane and ethyl acetateYield:- 76%
O
O
O
O
diphenyl ((3a,4,5,6,7,7a-hexahydro-1H-indol-3-yl)(hydroxy)methyl)phosphonate
MP: 125-127oC
THIN LAYER CHROMATOGRAPHY (TLC)
WASHING OF IMPURE SYNTHESIZED COMPOUNDS
SPECTROSCOPIC STUDIES
ANTIBACTERIAL ACTIVITY
TLC
Hexane
Ethyl acetate
ANTICANCER ACTIVITY
EIGHT SYNTHESIZED COMPOUNDS
IR SPECTRA RESULTS FOR:
COMPOUND 1
C P O
OH
H
Cl
Cl
O
O
COMPOUND 8
O
O
C
H
OH
P O
O
O
COMPOUND 9
ANTIBACTERIAL ACTIVITY
MICROBOTH DILUTION METHOD WAS USED TO TEST THE MINIMUM INHIBITORY CONCENTRATION AND MINIMUM BACTERICIDAL CONCENTRATION OF THE SYNTHESIZED COMPOUNDS
Staphylococcus aureasATCC-25925
Bacillus cereusATCC 10702
Vibrio fuvialisAL004
Escherichia coliATCC 10819
MINIMUM INHIBITORY CONCENTRATION (MIC)
Table 1: Antibacterial activities of 8 novel synthesized alpha-Hydroxyphosphonates.
NOTE: ---- denotes the resistance towards the test antibacterial agent at a maximum test concentration of 5mg/ml.
Microorganisms Synthesized compounds
(alpha-Hydroxyphosphonates)
Controls
C1 C2 C3 C4 C5 C6 C7 C8 Ciprofloxacin (+)
water (-)
Staphylococcus
Aureus ATCC-25925
5mg/ml
----
----
5mg/ml
----
----
----
----
0.625µg/ml
na
Vibrio fluvialis AL004
5mg/ml
----
5mg/ml
5mg/ml
5mg/ml
----
5mg/ml
5mg/ml
2.5µg/ml
na
Bacillus cereus ATCC 10702
5mg/ml
----
----
----
----
----
5mg/ml
----
0.313µg/ml
na
Escherichia coli ATCC 10819
5mg/ml
----
----
----
----
----
5mg/ml
----
0.625µg/ml
na
MINIMUM BACTERICIDAL CONCENTRATION (MBC)
Table 2: MBC results for the novel alpha-Hydroxyphosphonates
Microorganisms
Compounds
Control(+) C1 C3 C4 C5 C7 C8 Ciprofloxacin
Staphylococcus
aureus ATCC-25925
> 5mg/ml
5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5µg/ml
Vibrio fluvialis AL004
> 5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5µg/ml
Bacillus cereus ATCC 10702
> 5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5µg/ml
Escherichia coli ATCC 10819
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
>5mg/ml
5µg/ml
ANTICANCER ACTIVITY
The discovery of lung cancer and other cancers can synthesize and secrete acetylcholine by these cancers (Song and Spindel, 2008).
The inhibition of the enzyme Acetylcholinesterase can decrease the risk of the development of cancer.
The anticancer activity was determined using the Acetylcholinesterase Assay Kit (MAK119).
AChE Activity (units/L) = (A412)final – (A412)initial x 200
(A412)calibrator – (A412)blank
ACETYLCHOLINESTERASE ACTIVITY
Table 1: The results showing the activity of Acetylcholinesterase at 1mg/ml of the compound (in 100 % DMSO).
Compound Acetylcholinesterase
activity
1 -33.63
2 -49.78
3 +14.80
4 +14.57
5 +9.64
6 -19.73
7 -81.84
8 -22.87
KEY:- No activity+ Activity
TABLE 2: The results showing the Acetylcholinesterase activity at 0.1mg/ml of the compound ( in 100% methanol).
Compound Acetylcholinesterase
activity
1 -27.62
2 -27.11
6 74.68
7 -23.02
8 -134.53
CONCLUSION
A convenient high-yielding one-pot, two-component reaction of various aldehydes with
dialkylphosphite was successfully accomplished via Abramov reaction without any by-
products.
1,4-Dimethylpiperazine was proved to be an efficient catalyst in all the reactions.
The highlighting advantages of Abramov reaction.
The results of the antibacterial and anticancer activities revealed that the 8 compounds
screened had the bioactivities that were performed.
Further work from this project
To perform some more bioactivities such as the anti-fungal and antiviral activities of the newly synthesized compounds.
Acknowledgements
I would like to thank the following people: Prof A.R Opoku (supervisor)
: Dr C Sampath (Co-Supervisor)
: Dr R.A Mosa and Dr D Penduka (f (for helping out with bioactivities)
REFERENCESK. B. Dillon, F. Mathey and J. F. Nixon, Phosphorus. The Carbon Copy, John Wiley. & Sons, New York, 1998.
Song Pingfang and Eliot R. Spindel (2008), Basic and clinical aspects of non-neuronal Acetylcholine: expression
of non-neuronal acetylcholine lung cancer for therapy, Journal of pharmalogical sciences, (106): 180-185.
Todd, Science, 127,787, 1958.
Reddy S. Subba, Ch. SyamaSundar, S. Siva Prasad, E. Dadapeer, C. Naga Raju and C. Suresh Reddy. (2012), Synthesis, spectral characterization and antimicrobial activity of α-hydroxyphosphonates, Der Pharma Chemica, 4(6):2208-2213.
O. M. Friedman, E. Boger, V. Grubliauskas and H. Sommer, J. Med. Chem., 6, 50, 1963.
A. N. Pudovik, I. V. Konovalova, Synthesis, 81, 1979.
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