This document summarizes the synthesis and characterization of novel organophosphorus compounds and their biological evaluation. Eight α-hydroxyphosphonate compounds were synthesized using an Abramov reaction involving various aldehydes and diphenyl phosphite. The compounds were characterized using techniques like IR spectroscopy and melting point determination. Their antibacterial activity was tested against four bacterial strains using MIC and MBC assays, with some compounds showing activity. The anticancer potential was evaluated using an acetylcholinesterase inhibition assay, with some compounds exhibiting inhibition of the enzyme. In conclusion, a convenient method for synthesizing α-hydroxyphosphonates was developed and some of the compounds demonstrated selected biological activities.

1. 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

2. 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.

3. APPLICATIONS
 VERTERINARY MEDICININE PHARMACEUTICALS
 AGRICULTURE - INSECTICIDES

4. INTRODUCTION TO ALPHA-HYDROXYPHOSPHONATES
What are α-Hydroxyphosphonates?
α- Hydroxyphosphonates are class of Organophosphorus compounds
of synthetic interest because of their biological activity.

5. 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)

6. 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 α-Hydroxyphosphonates
(Subba et al., 2012).

7. AIM OF THE CURRENT RESEARCH PROJECT
To synthesize Novel α-Hydroxyphosphonates and test their
anti-bacterial and anti-cancer activities.

8. 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 anti-bacterial and anti-cancer activity of the synthesized
compounds.

9. MATERIALS AND METHODS

10. RESULTS AND DISCUSSION
.
MP: 80-82oC
COMPOUND 1
+ PH
O2N C P O
1,4-Dimethyl piperizine
THF (680C), Reflux 4-5 hours
OH
H
O
4- Nitrobenzaldehyde Diphenyl phosphite
TLC system:- 7:3 Hexane and ethyl acetate
Yield:- 80%
O
O
Diphenyl (hydroxy(4-
nitrocyclohexyl)methyl)phosphonate
O O
O2N CHO

11. MP: 74-76oC

12. MP: 130-132oC
COMPOUND 3
N
CHO
+ PH
O
C HHO
P
O
1,4-Dimethly piperizine
THF (680C), Reflux 4-5 hours
4-pyridinebenzaldehyde Diphenyl phosphite
TLC system:- 7:3 Ethyl acetate and hexane (suitable system)
7:3 Hexane and ethyl acetate (not suitable system)
Yield:- 78%
O
O
diphenyl hydroxy(pyridin-4-
yl)methylphosphonate
OO
N

13. MP: 60-69oC
+ H
P
O
C
OH
H
P
3-Pyridine aldehyde Diphenyl phosphite
COMPOUND 4
1,4-Dimethyl piperizine
THF (680C), Reflux 4-5 hours
TLC system:- 7:3 Hexane and ethyl acetate
O
O O
O
O
Diphenyl (hydroxy(piperidin-3-
yl)methyl)phosphonate
N
CHO
N
Yield: 76%

14. MP: 73-75oC

16. MP: 125-127oC

17. THIN LAYER CHROMATOGRAPHY (TLC)
WASHING OF IMPURE SYNTHESIZED
COMPOUNDS
SPECTROSCOPIC
STUDIES
ANTIBACTERIAL ACTIVITY
TLC
Hexane
Ethyl acetate
ANTICANCER ACTIVITY
EIGHT SYNTHESIZED COMPOUNDS

18. IR SPECTRA RESULTS FOR:
COMPOUND 1

19. C P O
OH
H
Cl
Cl
O
O
COMPOUND 8

20. O
O
C
H
OH
P O
O
O
COMPOUND 9

21. ANTIBACTERIAL ACTIVITY
MICROBOTH DILUTION METHOD WAS USED TO TEST THE MINIMUM INHIBITORY CONCENTRATION AND
MINIMUM BACTERICIDAL CONCENTRATION OF THE SYNTHESIZED COMPOUNDS

22. Staphylococcus aureas
ATCC-25925
Bacillus cereus
ATCC 10702
Vibrio fuvialis
AL004
Escherichia coli
ATCC 10819

23. 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

24. 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

25. ANTICANCER ACTIVITY
The discovery of lung cancer and other cancers , can synthesize and
secrete acetylcholine by these cancers (Song and Spindel, 2008).
Acetylcholine Acetate + Choline
The anticancer activity was determined using the
Acetylcholinesterase Assay Kit (MAK119).
AChEActivity(units/L)=(A412)final–(A412)initial x200
(A412)calibrator–(A412)blank
Acetylcholinesterase

26. 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

27. 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

28. CONCLUSION
 A convenient high-yielding one-pot, two-component reaction of various aldehydes with
diphenylphosphite 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 are
1) Shorter reaction times
2) Simple experimental procedure
3) Simple purification method
 The synthesized compounds were found to be having the selected biological activities.

29. FUTURE WORK FROM THIS PROJECT
To synthesize novel α-Hydroxyphosphonates containing
heteroatoms and to study their biological activities.

30. Acknowledgements
I sincerely 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)

31. REFERENCES
1) K. B. Dillon, F. Mathey and J. F. Nixon, Phosphorus. The Carbon Copy, John Wiley. & Sons, New York,
1998.
2) 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.
3) Todd, Science, 127,787, 1958.
4) 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.
5) O. M. Friedman, E. Boger, V. Grubliauskas and H. Sommer, J. Med. Chem., 6, 50, 1963.
6) A. N. Pudovik, I. V. Konovalova, Synthesis, 81, 1979.