1. Not so Apple-Licious: The Reduction of Methylglyoxal Using Granny Smith, Golden
Delicious and Macintosh Apples
Jennifer Driskell, Jennifer Falkowski, Elizabeth Reed and Chelsea Ruston
The Pennsylvania State University, Chemistry Department Course 213 B
Introduction Results Conclusion
Background: Figure 2: IR Analysis Methylglyoxal Figure 3: NMR Analysis Methylglyoxal The methylglyoxal standard did not produce conclusive results, which could contributed
•The commercial process for making artificial sweeteners (sugar alcohols) involves to its improper temperature storage conditions
importing the raw materials, such as glucose. The glucose is hydrogenated using a nickel The methylglyoxal produced an alcohol stretch on the IR and did not produce an
catalyst to produce sorbitol. Further filtering and purification is required to isolate and aldehyde peak on the NMR indicating that the sample could have been partially oxidized
refine the sorbitol for use as an artificial sweetener.1 The expected alcohol and C-H stretches for propylene glycol were observed via IR, while
•Artificial sweeteners provide many advantages to traditional sugars in that they have the starting material’s ketone and aldehyde stretches were not observed. The apples did
lower caloric value than sucrose and are shown to reduce the incidence of dental caries.1 not reduce all of the methylglyoxal into propylene glycol producing a mixture of four
Goal: products shown in figure 15
The goal of this experiment was to use the enzymes present within the apple to reduce The NMR analysis showed that the aldehyde peak was still present for each of the apple
the compound methylglyoxal to propylene glycol. preparations which indicate the enzyme mediated reduction did not completely reduce
Hypothesis: Figure 4: IR Analysis Propylene Glycol Figure 5: NMR Analysis Propylene Glycol the aldehyde of methylglyoxal
•It was hypothesized that from the three apple preparations (mashed, sliced and peel) The peaks of interest on the NMR are hard to differentiate due to the possibility of
the sliced would produce the best yield of propylene glycol because the reduction would many forms of reduction from the starting material
required intact cells to produced the enzymes needed to reduce the ketone and Alcohol groups were observed between 3600-3200 cm-1 and ketone and aldehyde
aldehyde to alcohol groups.2 stretches were observed around 1660 and 1715 cm-1 in the IR analysis indicating that a
•It was thought that the colored apple peel would influence the enzymes present in the mixture of reductions took place using the apple enzymes
apple and thus the ability to reduce methylglyoxal. Scientific research has shown that The TLC analysis using the Vanillin and DNP dyes both confirmed that ketone and
specific enzyme activity varies in apples depending on which season they ripen. Since the aldehyde groups were present within the product, however, the Vanillin dye also
apples that are being utilized all have different peak ripening seasons it is theorized that suggested that there were trace amounts of alcohol groups present
one of them will have a superior reducing ability compared to the rest.3 The apple preparation that produced the best product was mashed with a 68 percent
Relevance: Figure 6: IR Analysis Green Mashed Figure 7: IR Analysis Green Sliced yield, it was also observed that the peel produced the worst amount of product possibly
This experiment would provide a greener way to produce sugar alcohols by using apples because the peel would not contain enzymes needed for the reduction
as the reducing agents instead of chemicals and/or bacteria. These naturally produced
sugar alcohols would potentially be safer and cheaper than the chemically made
sweeteners on the market. Figure 15: Reduction Possibilities of Methylglyoxal
Figure 1 Reduction of methylglyoxal to propylene glycol
O O OH
OH
H H
H H H
H3C Apples
H
+ H3C + H3C
H3C
Figure 8: IR Analysis Green Peel Figure 9: NMR Analysis Green Mashed H
Water H OH OH
O O
Methylglyoxal Propylene Glycol Acetol Lactaldehyde
Materials
Granny Smith Apple Vanillin
Golden Delicious Apple 2,4-Dinitrophenylhydrazine (DNP)
Macintosh Apple Sodium Borohydride
Apple Peeler Further Experimentation
Methylglyoxal Instruments Devise better extraction techniques to remove all of the propylene glycol from the
Figure 10: NMR Analysis Green Sliced Figure 11: NMR Analysis Green Peel resulting solution
Distilled Water  400 MHz 1HNMR
Ethyl Acetate 60 MHz 1HNMR Use a simpler model system that contains only one carbonyl group
Ethanol  Infrared Spectrometer Store compounds in proper storage conditions
Dichloromethane Orbital Shaker Look for products that will result in crystal structures instead of an oil/wax
TLC Plates Use a polarimeter to determine the stereospecificity of the obtained product
Perform a reduction using the reducing agent, Lithium Borohydride
Run GC and GC/MS on the product to determine actual structures
Procedure Have only one variable that is being tested
Procedure 1: Try other fruits or vegetables that contain a stereospecific reducing enzyme
Apples were rinsed using distilled water Figure 12: Vanillin TLC Dye Figure 13: DNP TLC Dye
Three different apple preparations were used: mashed, slices and the peel
Approximately 2 grams of each apple preparation, 50 mL of distilled water and 1 mL of
Yellow Red Green Standard
methylglyoxal were added to individual Erlenmeyer flask Standard Yellow Red Green References
The flasks were incubated at 30 C on an orbital shaker set at 150 rotations per minute 1 Nabors, Lyn O’Brien. Alternative Sweeteners. New York: Marcel Dekker, Inc., 2001.
for approximately 50 hours 2 Bohman, Bjorn; Cavonius, L.R.,; Unelius, C. Rikard. The Journal of Green Chemistry, 11
The solution was separated from the apples and liquid –liquid extraction was used to ed. 2009: 1900-1905.
isolate the propylene glycol product 3 Simonov, I. N. Biochemical and Biological Differences Between Apple Varieties. Trudy
The organic layer containing ethyl acetate and the product propylene glycol was dried Moskovskoi ordena Lenina Sel’skokhozjaistevennoi Akademii imeni K.A.
under a flow of nitrogen gas Timirjazeva, 1940 Vol 4, pp 71-101.
The weight of the product was obtained once the product was completely dry
The samples were prepared for NMR and IR analysis Figure 14: Percent Yield for Green Apple
Procedure 2: 80
68
Thin-layer chromatography was used to separate the compounds and then dyes were
70
60
applied to view the spots Acknowledgements
Percent Yield (%)
50
A standard for each dye was developed spotting only methylglyoxal and propylene 40
glycol on the TLC plate 30 The Pennsylvania State University Chemistry Department
20
The product of each apple reaction was re-suspended using a small amount of 10 5
15
Course Coordinator: Dr. Sheryl Rummel
dichloromethane (DCM) and then TLC and dyes, vanillin and 0
Green Peel Green Sliced Green Mashed
Teaching Assistant: Brad Landgraf
dinitrophenylhyrazine, were used to predict the presence of ketone/aldehyde and Apple Preparation The Instrument Room TAs
alcohol groups