Determining the pH Effect and Partition Coefficient of Diethyl Malonate in Water and Water - Pentane systems using Liquid-Liquid

Extraction and Ultraviolet - Visible Spectroscopy.

Chris BilhamAmanda Ranero

Dr. Rainer VolkamerDr. Molly LarsenZachary FinewaxTheodore Koenig

University of Colorado Department of Chemistry

and Biochemistry

❖Agenda

• Introduction• Motivation• Methodology • Results & Analysis• Conclusions• Recommendations• Acknowledgements

diethyl malonate

Image from: http://www.sigmaaldrich.com/catalog/product/aldrich/d97754?lang=en&region=US

❖Introduction to Diethyl Malonate (DEM)

What is Diethyl Malonate?• Diethyl Malonate (DEM) is the diethyl ester

of malonic acid.

• DEM is found naturally in grapes and strawberries

• It is a colorless liquid with an odor like apples (used in artificial apple juice)

• DEM is favorable for use in industry because it is the most stable form of malonic acids/esters

Diethyl Malonate

Malonic Acid

Diethyl Esters

https://en.wikipedia.org/wiki/Malonic_acidhttps://en.wikipedia.org/wiki/Diethyl_malonate

Major Uses of DEM

● Pharmaceuticals/Chemical Precursor (Biggest Market Value)● Fragrances/Perfumes● Dyes● Adhesives● Artificial flavorings

❖Introduction to Diethyl Malonate (DEM)

Current Method of DEM Production

● Current method uses toxic and environmentally hazardous materials

● Use of such materials not only is detrimental but incurs high waste disposal costs

http://www.directindustry.com/prod/solaris-biotechnology/product-54387-443440.html https://en.wikipedia.org/wiki/Malonic_acidhttp://culmotrialattorneys.com/bpa-may-alter-gene-regulation/

Motivation for Investigation

● All senior chemical engineer undergraduates are required to complete a design project for an assigned client.

● Our client DMC limited has produced an environmentally friendly way to produce DEM using engineered microbes and a sugar feedstock.

● However the majority of DEM produced is done in organic solvents and there was little to none literature on how to separate DEM from an aqueous solution.

● Our design team proposed two continuous separation methods: 1) Liquid-Liquid Extraction (LLE) with distillation using n-pentane. 2) Natural Phase Separation with decanting.

● Both the above methods were missing critical physical parameters in order to develop a successful process model. Mainly, the partition coefficient for our solvent n-pentane and the effect of pH on the solubility of DEM in water.

❖Motivation

Depiction of Separation Process Series Process of Phase Separation followed by LLE and Distillation

.

❖Motivation

Critical for these Units

❖Methodology

Choice of Method of Quantification

• It was determined (by recommendation of Theodore Koenig) that the best method of quantification would be to use Ultraviolet – Visible Spectroscopy (UV-VIS)

• Originally High Performance Liquid Chromatography coupled with UV-VIS spectroscopy and Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy (ATR - FTIR) were considered, but subsequently discarded.

• It was found that DEM shows an absorbance peak at 212 nm, in the Ultraviolet region, using a Cary 5000 UV-VIS spectrometer.

• It was found that n-pentane and water had a cut off absorbance around 190 nm, preventing spectral interference

Figure 1: UV-VIS data from Cary 5000

❖Methodology

Instrumentation

Cary 5000 UV-VIS spectrometer • 1 cm path length quartz cells• 0.5 nm data interval• 1 nm slit width• 200nm – 350 nm scan range

125 mL separatory funnel 60 mL separatory funnel

https://nfm.kaust.edu.sa/PublishingImages/Cary_5000.jpg

❖MethodologyExperiment 1: pH Effect on DEM solubility on water

• 50 mL of solutions at a pH of: 3.92, 4.92, 5.92 and 7 were prepared from 12 M Hydrochloric Acid.

• A magnetic stir bar was added and set to run around 7-8 (rigorous stirring)

• DEM was titrated into each solution until visible phase separation was observed.

Experiment 2: Determining the partition coefficient for DEM in water/pentane systems, with pH affect

• Calibration curves were produced for DEM in both n-pentane and water using 5 standards and three or more replicates

• A 0.0662 ± 0.0004 M (approx. 10630 ± 60 ppmm) standard aqueous solution was used for all aqueous trials.

• Two trials were done at a pH of 4.92 and 5.92.• Extractions were done at 1:1, 1:7.5 and 1:10 ratios of solvent

to water. • Heavy Phase and Organic phase were collected, diluted into

appropriate linear range and quantified with the Cary 5000 UV-VIS

Table 2: Extraction Volumes

pHVolume

Water(mL)Volume

Pentane (mL)Neutral 50 50Neutral 50 50Neutral 50 5Neutral 75 5

5.92 50 54.92 100 10

Conditions at 22 °C, 1 atm

Diethyl Malonate

Pentane

Boiling Point (°C) 199 36

Melting Point (°C) -50 -130

Molecular Weight (g/mol)

160.17 72.15

Density (g/mL) 1.05 0.626pKa 12.9 45

Table 1: Physical Data

❖Experiment – 1 Results & Analysis

Results: pH effect of solubility of DEM in water

• It was found that lowering the pH did decrease the solubility of DEM in water

• The scale was done as a relative solubility in order to standardize the ‘limit’ of solubility.

• Although the exact mechanism is not clear, we hypothesis that the increase in [H+] ions is acting to make the solution ‘more polar’ driving DEM out of solution.

• This effect would be similar to that of salting out, increasing hydrophobic interactions with DEM.

• Although terms are not known, equation is shown to demonstrate it is a non-linear affect.

% 𝑅𝑆=[𝐷𝐸𝑀 ]𝑝𝐻

[𝐷𝐸𝑀 ]𝑁𝑒𝑢𝑡𝑟𝑎𝑙×100

Figure 2: pH affect on relative solubility of DEM in water

❖Experiment – 2 Results & Analysis

Calibration Curves for DEM in Water and n-Pentane Respectively

Figure 3: Calibration Curve for DEM in water Figure 4: Calibration Curve for DEM in n-pentane

❖Experiment – 2 Results & Analysis

Molar Absorption Coefficient (σ)

Figure 3: Molar Absorption Coefficient Epsilon in inverse (Molarity*cm)

σ=  𝑨𝒄𝒍

Beer- Lambert Laws Solved for Molar Absorption Coefficient

pHVolume

Water(mL)Volume Pentane

(mL)Parition

Coefficient KError

Neutral 50 50 2.797 0.08Neutral 50 50 2.627 0.09Neutral 50 5 2.529 0.07Neutral 75 5 5.344 0.110

5.92 50 5 2.514 0.074.92 100 10 2.756 0.1

Mean 2.645 0.080STDV 0.129LOD 0.00304LOQ 0.00024 pH

Volume Water(mL)

Volume Pentane (mL)

Parition Coefficient K

Error

Neutral 50 50 12.121 0.7Neutral 50 50 9.525 0.7Neutral 50 5 3.820 0.3Neutral 75 5 3.172 0.3

5.92 50 5 4.810 0.44.92 100 10 4.815 0.4

Mean 6.377 0.4STDV 3.595LOD 0.0791LOQ 0.0058

❖Experiment – 2 Results & Analysis

Partition Coefficients Determined from Liquid-Liquid Extractions

Table 4: Partition Coefficients from Aqueous (Heavy Phase)

Table 5: Partition Coefficients from Pentane (Light Phase)

• A two sided t-test on the means concluded these values were statistically

different at the 95% confidence level• A 95% confidence interval for pentane and

water respectively are: 6 ± 6 & 2.6 ± 0.2

𝐾=𝑉 𝑎𝑞

𝑉 𝑜𝑟𝑔𝑞𝑎𝑞−𝑉 𝑎𝑞

𝑉 𝑜𝑟𝑔

𝐾=𝑉 𝑎𝑞

𝑉 𝑜𝑟𝑔( 1𝑞𝑎𝑞

−1)Ratios can ‘blow up’ results

❖Experiment – 2 Results & Analysis

So why did that happen?

• The main contribution is hypothesized to be the evaporation of pentane, causing a variable increase in concentration.

• This is further validated by the fact that pentane samples were not run on the same day as the LLE; K values for samples run the longest time after LLE were the highest.

• It is also hypothesized that a micro emulsion was forming with DEM saturated water. Pentane with the smallest LLE volume would be most susceptible to these effects while the large volume of aqueous phase could be sampled while ignoring this emulsion.

??

❖Experiment – 2 Results & Analysis

pH and Partition Coefficient Analysis

2.645𝐾 𝑝𝐻=

𝐾 [𝐻 ]𝐾 𝑎+[𝐻 ]

• The given equations below are used to describe the theoretical change in K based upon a known K and pH

• This data was graphed to the right as figure 5

• Looking at the pH range for this experiment it is expected that K did not change in our pH range. Our findings match the theoretical.

Figure 5: Theoretical effect of pH on K; pka = 12.9, Ko = 2.645

❖Conclusion

• pH does decrease the solubility of DEM in water, the effect is non-linear and most likely operates in the same manner as ‘salting out’.

• The partition coefficient was found to not be dependent on pH in our experimental range, this matches theoretical predictions.

• The partition coefficient of DEM in water-pentane systems was found at a 95% confidence interval to be 2.6 ± 0.2 from aqueous data and 6 ± 6 in the organic n-pentane phase. These results were found to be statistically different by a two sided t-test

• The pentane data is deemed to be precise but not accurate.

❖Recommendations

• A larger pH range should be investigated (1-14) with replicates to better examine effect of pH on DEM solubility in water.

• Additionally salt concentrations should be done in a similar manner to verify pH effect hypothesis and to further characterize DEM-water solubility

• A higher pH range should be investigated (7 - 14) with replicates to better examine effect of pH the partition coefficient

• Larger separatory funnels should be used, allowing for larger aqueous and pentane volumes to mitigate the effect of emulsion layers

• Pentane samples should be either be run immediately after or stored in a manner that prevents evaporation or can be accurately re-diluted

❖Acknowledgements

Professor Rainer Volkamer

Dr. Molly Larsen

Zachary Finewax Theodore Koenig

University of Colorado Department of Chemistry

and Biochemistry

❖References

Acid-Base Equilibria. (n.d.). Retrieved April 25, 2016, from http://chemed.chem.purdue.edu/genchem/topicreview/bp/ch17/ph.php

Harris, Daniel C. "Fundamentals of Spectrophotometry." Quantitive Chemical Analysis. 8th ed. China Lake: W.H. Freeman, 2010. 505-565. Print.

Larsen, Molly, and Rainer Volkamer. "E6 Analysis of Vanillin in Vanilla Using High Performance Liquid Chromatography." Chem 4171/4181 Instrumental Analysis Laboratory. Fall 2015 ed. 75-79. Print.

Lipscomb, M., Dr. (2016). Client Meetings [Personal interview].

Dr. Matthew Lipscomb, PhD in Chemical Engineering from CU Boulder. Client for DMC limited.

S. J., Dr. (2005, April 11). MALONIC ACID DIESTERS Dimethylmalonate, 108-59-8 Diethylmalonate, 105-53-3. Retrieved March 15, 2016, from http://www.inchem.org/documents/sids/sids/malonates.pdf

Used, extensively throughout the report

Method for Producing Diethyl Malonate. (2013). Retrieved April 25, 2016, from https://xudaphne01.wordpress.com/2013/08/28/method-for-producing-diethyl-malonate/