(490g) Analysis of Biodiesel: Use of HPLC with An ELSD, Size Exclusion Method

Introduction

            Recent discoveries and growth in the field of Biodiesel synthesis has created a necessity to use fast, accurate, and easy methods for the analysis of the final product.  The products and byproducts of the synthesis of Biodiesel include Biodiesel (FAME), triglycerides, diglycerides, monoglycerides, and glycerin.  The most common analysis method used is to analyze the Biodiesel phase, which should include the tri-, di-, and mono- glycerides as well as the Biodiesel.  By creating the derivative of the sample, it is possible to then evaluate the conversion of Biodiesel in a GC column as described by Monteiro et al. [2].  This process requires the integration of multiple peaks per component, and comparing the amounts of the glycerides with the amount of Biodiesel found from the resulting signal output.

It has also been observed that over time the column tends become less accurate and can also introduce false readings as well as the problem of baseline drifting occurring as stated by Monteiro et al. [2].  The use of a HPLC with ELSD was then employed to compare to the analysis methods of a GC procedure.

                Analyzing the Biodiesel in a HPLC with ELSD is a very easy method.  Unlike a GC column, the HPLC does not require the derivatives to be produced in order to analyze in the column.  The HPLC only requires a dilution with a volatile liquid mobile phase in which the products are soluble.

Materials and Methods

                The methods for creating the Biodiesel were performed by using microwave catalysis as described in papers by Tompsett et al. [3] and Leadbeater et al. [1].  A heterogeneous catalyst of sodium aluminate was used to ensure that the solid catalyst would not be dissolved in the sample as a homogenous catalyst may become.

                An EnSight? Biodiesel Analyzer provided by W.R. Grace was employed as the HPLC with ELSD used for analyzing the Biodiesel.  With the size exclusion column setup in the HPLC, mobile phases of ethyl acetate and methanol were utilized.  The mobile phases are employed to pump the sample through the column.  The dilution was then performed with either one of the two mobile phases being used.  Dilutions ratios were experimented to determine the best dilution for the most clear and readable graph.

                Different ratios of mobile phase flow rates were used, keeping the overall flow rate constant at 0.8 mL/min.  Ratios of 85%, 50%, and 15% of ethyl acetate were tested to determine the best method for analyzing the samples (remaining parts were methanol).  The Biodiesel and glycerin phases were analyzed separately as well as in the same run.

Results and Discussion

                The Biodiesel layer was analyzed, as this layer contains the undesired mono-, di-, and tri- glycerides, as well as the desired FAME components.  Many dilution factors were experimented with, and it has been observed that a dilution of 750:1 is an optimal dilution to be analyzing with.  At dilution factor less that 750 parts of the mobile phase, double peaks were observed in the results. Ethyl acetate was used in the dilution of the Biodiesel phase, along with the mobile phases running at 85% ethyl acetate and 15% methanol. By analyzing the Biodiesel phase first, the triglyceride peak was observed at roughly 7.8 minutes, the diglyceride peak at roughly 8.2 minutes, and the Biodiesel peak at roughly 8.8 minutes.  The monoglyceride peak was not observed in this phase, contrary to previous work.

                After analyzing the glycerin phase, also at a 750:1 dilution with ethyl acetate, it was observed that the monoglycerides were present in this phase; however, the glycerin was not being detected by the ELSD.  Upon further analysis, methanol was then used to dilute the glycerin phase, as the glycerin was more soluble than in the other mobile phase.  Manipulating the ratio of flow rates of the mobile phase, it was determined that by diluting with methanol and running the flow at 85% methanol, a glycerin peak was observed at roughly 11.3 minutes.

                Using the results and methods from analyzing the two phases individually, the two phases were then combined with a dilution in a 50:50 mixture of ethyl acetate and methanol.  The mobile phases were then run at 85% methanol and 15% ethyl acetate.  The sample that was analyzed was at a ratio of 9.8:1 Biodiesel to glycerin, as this was determined from the volumes of each phase of the product.  The results clearly display the presence of all five peaks, seen in figure 1.   The final two peaks are difficult to see, as they are very small compared to the first three area peaks.  Figure 2 below is zoomed in to show that the final two peaks are present.

Figure 1. Resulting chromatograph from a 13 minute run with a 750:1 dilution of an ethyl acetate and methanol mix (50:50) to a Biodiesel and glycerin (9.8:1) mix.  The resulting peaks exit in the order of tri- (7.9 min), di- (8.3 min), Biodiesel (8.8 min), mono- (9.3 min), and glycerin (11.3 min).  The final two peaks can be seen clearly in figure 2.

Figure 2. Resulting chromatograph from a 13 minute run with a 750:1 dilution of an ethyl acetate and methanol mix (50:50) to a Biodiesel and glycerin (9.8:1) mix, zoomed in over the timeframe where the monoglyceride (9.3 min) and glycerin (11.3 min) peaks were observed.  These peaks are much smaller than the first three, which is expected as there is much less of them present in the sample.

Significance

The use of a HPLC with ELSD is a new analysis method for the conversion of Biodiesel.  The ability to not only have each of the components represented by their own single peak, but also the ability to analyze both the Biodiesel and glycerin phases simultaneously is a great advantage in time and resources to the GC analysis methods.

References

1.      Leadbeater NE, Stencel LM (2006) Energy & Fuels 20: 2281

2.      Monteiro, Marcos Roberto, Alessandra Regina Pepe Ambrozin, Luciano Morais Liao, Antonio Gilberto Ferreira. Talanta 77 (2008) 593-605.

3.      Tompsett, Geoffrey, et al, "Heterogeneous Catalysis of Biodiesel: Rapid and Selective Microwave Synthesis." Department of Chemical Engineering, University of Massachusetts Amherst

Acknowledgement to: W.R. Grace for providing the EnSight? Biodiesel Analyzer