1. Abstract
Nickel is not a common element naturally available due to its reactivity with oxygen; however it
serves an important role as the metal center for a metalloenzyme, Nickel Acireductone Dioxygenase (Ni-
ARD),found in the methionine salvage pathway. Ni-ARD, one of the only dioxygenases containing
nickel, is a critical component of a recycling pathway in which the amino acid methionine is
recycled in bacteria and plants. There are multiple enzymes present throughout the pathway, but
the cause for the reaction with the substrate acireductone resulting from Ni-ARD versus Fe-
ARD, whose weights only differ from which metal center is in place, is the area focused on in
this research. In the methionine salvage pathway the Fe-ARD acts to continue the pathway, to
allow methionine to be recycled, and the other associated reactions to continue to occur within
the pathway. These associated reactions are involved in the synthesis of polyamines and
ethylene, required for cell growth in bacteria and fruit ripening in plants, to continue. However,
Ni-ARD acts to discontinue the pathway not allowing methionine to be recycled, and preventing
the other associated reactions from taking place. This stark difference in function of the two
ARD enzymes causes speculation as to what is actually occurring at their active sites, where the
metal and surrounding structures bind and interact with the ligands. To determine how the metal
center is reacting with the surrounding protein based ligands would allow an understanding of
why Ni-ARD interacts with the substrate, acireductone, in a way to discontinue the pathway.
The structure and function of the enzyme are determined partially by the center’s active site. This
is the basis for a novel family of metal complexes that have been synthesized and characterized
to model the structural and electronic properties at the Ni-ARD active site. Zinc and Nickel
analogous metal complexes with slight structural alterations were used for comparison to begin
analyzing the structure-function relationship of these models.
The process of biomimetic modeling will be utilized to create chemical models in the lab to
mimic the chemical environment found at the active site of the Ni-ARD enzyme. By creating a
family of eight proposed metal complexes, each comprised of a slight variation in structure,
analysis and later reactivity tests, will allow examination of the structure function relationship
witnessed between the metal and surrounding ligand. Nickel being reactive with oxygen and
paramagnetic due to unpaired electrons in its d-orbital, requires the use of Zinc to develop
methods along with characterization. Zinc analogues of the metal complexes are first created to
develop methods to give a basis to follow for the synthesis of the Nickel complexes.
This paper will discuss the progress made so far on the production of the assigned Zn and Ni
complexes; [ZnII(OHMe26-H-DPPN)](OTf)2, and [NiII(OHMe26-H-DPPN)](OTf)2.

2. Experimental section
Synthesis of 1-(tert-Butyloxycarbonyl)propyl diamine)(NPNBoc)
Di-tert-butyl-dicarbonate (Boc) (3.05 g, 13.9 mmol) was dissolved in 200ml of dichloromethane
into a 250 ml beaker. The solution was placed into the addition funnel. Separately 1,3-
diaminopropane (7.05 ml, 83.8 mmol) was dissolved in 25 ml of dichloromethane and placed in
a 500ml round bottom flask with a fitting that matches the addition funnel. A stir bar is also
added to this round bottom flask. The Boc solution was added to the ethylene diamine solution
in the round bottom flask at room temperature dropwise via addition funnel over 6hr and allowed
to stir overnight. This reaction should be monitored to ensure that dripping continues steadily for 6hrs,
at a rate of approximately 1 drop/sec. At the end of the overnight process the contents from the round
bottom flask were filtered via gravity filtration and poured into a clean Erlenmeyer flask to remove the
white precipitate. Once everything was poured out of the round bottom flask ~10mL of dichloromethane
were used to rinse the inside of the flask to ensure transfer of all material. The resulting clear
dichloromethane solution containing the desired compound was added to a 500 ml separation funnel.
Following, ~200ml of a saturated sodium carbonate solution were added to the separation funnel,
resulting in a formation of 2 layers. The dichloromethane (organic) layer will be the one at the bottom due
to its density. A standard wash was performed on this mixture, involving sodium carbonate and sodium
chloride (to rid of unwanted ions). Organic layer is then drained into a clean 500ml conical flask. The
sodium carbonate “dirty” layer is poured into a separate ~500ml Erlenmeyer flask as well. The organic
dichloromethane layer is then put back on the separation funnel and the procedure is repeated one more
time by washing it with another ~200ml of saturated sodium carbonate. At the end of the second wash
there should be a combined dichloromethane organic layer from both of the washes and a combined
“dirty” sodium carbonate layer. The organic dichloromethane layer was added again to the separation
funnel and ~200 ml of brine were added to it on the separation funnel. This mixture was washed one more
time following the same process as that used for the sodium carbonate washes. The bottom organic
dichloromethane layer was collected into a clean Erlenmeyer flask and the brine solution was collected
into a separate,large Erlenmeyer flask. The combined organics were dried with Na2SO4 (enough to see no
more clumping of Na2SO4 ) to ensure removal of any extraneous water resulting from the washes. After
10-15 min of letting this solution stand the sodium sulfate drying agent is removed via gravity filtration
and the solution is placed into a pre-weighed 500ml round bottom flask. The solvent dichloromethane is
then removed via rotovap resulting in a clear oil. After the solvent is dried completely the weight of the
resulting flask is measured and the yield calculated for the reaction. Obtained 2 samples of NPNboc,
1.26g, and 2.04g of NPNBoc (51.9%, and 84% yield respectively).
Synthesis of N-[tert-Butyloxycarbonyl]-N',N'-[bis(2-pyridilmethyl)]propane-1,2-diamine (6-H-
DPPNBoc).
The synthesis of 6-H-DPPNBoc was adapted from the synthetic procedure described by Bernal et. al.
Both samples of NPNBoc (1.26g, and 2.04g) were dissolved, separately, in 5M NaOH (13.5 ml) and
added at room temperature to a solution of 2-picolyl chloride hydrochloride (2.37g, 3.84g) also dissolved
in NaOH (13.5ml). The reaction mixture was allowed to stir for three days at room temperature. At the
end of this period water (25 ml) was added to the mixture and this is extracted with dichloromethane (3x
50 ml) and the combined organics were washed with brine resulting in a pale orange oil. The oil was

3. purified via column chromatography (70:30 Acetone:DCM) to afford clean products (2.00g, 3.10g, and
43.8%, 67.8% yield).
Synthesis of [N,N-bis(2-pyridilmethyl)propane-1,2-diamine] (6-H-DPPN). 6-H-DPPNBoc (1.55 g) was
dissolved in dichloromethane (3.00 ml) at room temperature. 3.60 ml Trifluoroacetic acid (TFA) was
added and the brown mixture was allowed to warm up and stir at room temperature overnight. At the end
of this period all volatiles were evaporated and 5M NaOH (20 ml) added to the oily mixture. The aqueous
layer is extracted with dichloromethane (5x 50 ml) resulting in an orange oil the desired product in 98.4%
yield (1.27g).
Discussion and results
Sodium sulfate should be preferred drying agent unless more powerful one required (KOH).
Column conditions have shown to be optimum at a 70:30 (or 66:33, run TLC for comparison)
acetone:dcm ratio
Most challenges in drying of samples and compounds, due to problems with roto-vap and
vacuum pump (of shlank line).
Currently have 1 sample of 6-H DPPN ready for metal chemistry, and 2 samples of 6-H DPPN
Boc which need to be run through column and then deprotected before being ready for metal
complex synthesis.
NMR spectra to be taken when the project is picked up again during the fall

4. 6-H DPPN Boc

5. TLC results of post-column 6-H DPPN Boc showing successful purification
Conclusion
Learned how to work a shlank line.
Future work
Completion of synthesis and analysis of enzyme analogues.