Precipitate Solubility

Multiple times during the summer, a precipitate was observed when iron and curcumin were mixed.  During the past week the precipitate was isolated and dried on two occasions for testing.  The issue experienced in the summer remained problematic- the precipitate was not soluble in several of the common solutes that were tried.  Eventually, acid was used in an attempt to free iron from the precipitate to test if there was iron in the precipitate at all.  KSCN was added to the solution as the appearance of a red color would indicate that a FeSCN complex was being formed.

UV and HPLC were used to test the solution after it was heated with nitric acid in order to dissolve more of the precipitate.  Both UV and HPLC spectra showed the presence of the iron and curcumin complex that had previously only been confirmed in solution.

1/8/19
-Alex Griffith

Beginning of Winternship

Today, another reflux extraction of turmeric and methanol was performed in order to retest the curcumin and curcumin-iron complexes using HPLC.  A solution of curcumin and iron produced a precipitate that was filtered out and left to dry for further testing.  The filtrate was also saved.

The main problem experienced today was the malfunctions of the HPLC machine.  It is currently unclear what the issue was but no spectra could be run due to the pressure building too high and the machine subsequently shutting down.

1/2/2019
-Alex Griffith

Unknown Precipitate

To form the iron-curcumin complex, stoichiometric ratios are calculated and volumes of curcumin solution and iron solution are mixed.  Before the iron is added to the curcumin, the pH of the curcumin is adjusted to approximately 7.5 with triethylamine.  It was observed that when this solution is left overnight, a precipitate appears.  When centrifuged, it was discovered that the precipitate was actually a denser fluid and not a solid.  This dense fluid was dark in color and oily in consistency.  It was not soluble in the solvents typically used for FeCl₃ or curcumin.  After testing the oily fluid using HPLC it was discovered that it was not the iron-curcumin complex as had been reported in several papers, but just iron hydroxide.  

In order to further prove that the oily precipitate is not the iron-curcumin complex, different ratios of curcumin and iron(III) were mixed.  If the dense oil is iron hydroxide, then the test tubes containing more iron will have more precipitate.

7/31/18

-Alex Griffith

Trip to NJIT

On Wednesday, July, 25th, we went to NJIT to do mass spectrometry on a few samples of curcuminoids and curcuminoid complex we had prepared the day prior. We were greeted by Dr. Zhang, a research professor, and his post-doctoral assistant. They brought us to a Liquid Chromatography Mass Spectrometry machine to complete the experiments which would identify the masses of fragments of our samples.

We learned that, with the method used, the complex would not appear at its hypothesized molecular weight, and that the mass of one curcumin and one iron appeared to be the most stable fragment. An ongoing topic of interest is the phase, gaseous, or solvated, of the complex.

-Alexander Czechowicz

Microbiology Controls

This week marked an expansion of focus upon plating our hypothetical complex onto various TSA media containing various bacteria. Much of what we analyzed on this side of the project looked like the following:

We saw that the hypothetical complexes did not exhibit larger zones of inhibition across Al3+, Cu2+, Fe3+ and Fe2+ than curcumin alone. Dr. K hypothesized that the formation equilibrium had to be pushed to the right and we increased the mol:mol ratio of cation to curcumin for the next plates. This showed mild evidence supporting Dr. K's hypothesis.

We had more questions about achieving a high concentration of complex in solution and attempted to accomplish that goal by considering the time since mixing our curcumin and the cation. Dr. K had Isaac, who normally runs our HPLC, mix the solution in his syringe which produced results consistent with greater amounts of complex. We decided to try to apply this to the microbiology side of things.

Hypothetically, if we accept that the rate of decomposition of the complex is greater than its rate of formation shortly after mixing, then its concentration should be highest right when mixed. We will check back in next week to see whether or not the plates we streaked today support this hypothesis.

7/19/2018
-Alexander Czechowicz

Crystalization

After crystals were extracted from turmeric solution, it was noted that the crystals were not pure orange in color.  There were several very small flecks of yellow and red.  The lab supplied separated curcuminoids were all slightly different in color, one being yellow, another being orange, and the last being red.  From this information, it was hypothesized that if these crystals were separated by color, they could be used as seed crystals in a super saturated curcumin solution to produce larger single color crystals.  These single colors could possibly be the individual curcuminoids.  If true, this would mean there would be a much easier and cheaper way to separate curcuminoids.

Curcumin Crystals Before Separation

Separating the small crystals by hand had to be accomplished using a dissection microscope, a metal spatula, and tweezers.  Because the extremely low percentage of demethoxycurcumin and bisdemethoxycurcumin in turmeric, the amount of yellow and red crystals in the yield was very small.

Yellow, Orange, and Red Crystals: Seperated

7/18/18
-Alex Griffith

Melting Points

Several solids have been filtered from solutions containing both metal cations and curcumin.  Especially when it comes to iron(III), the identity of the filtered solid has been questionable.  It was possible that the solid was iron oxide and not the curcumin-iron complex.  As a method of identifying the solid, we attempted to find the melting point of the black crystals that were removed from solution.
The melting point of curcumin is 183°C.  The melting point of iron oxide is far beyond the limitations of the melting point apparatus at 1500°C.  The black crystals did not melt at 250°C, although they did begin to change color.  The limitations of the machine made further analysis of the black crystals impossible.

The melting point apparatus was also utilized to confirm the identity of orange crystals extracted from a turmeric solution.  

The melting point of the crystals was 179°C.  Because the melting points of the curcuminoids are all slightly different, the "impurity" caused by having all three curcuminoids together reduced the melting point, meaning that the crystals were confirmed as curcumin by the melting point testing.

7/12/18

-Alex Griffith