Qualitative Test for Conjugated Dienes in Eucalyptus Oil

Isabella Portillo, John Ferrara

Department of Chemistry, Binghamton University, Binghamton, NY 13902

02/23/2025
Abstract

In this experiment the focus is on the Diels- Alder reaction. This reaction occurs with

activated alkene with a conjugated diene. This type of reaction is very moisture sensitive, so the

goal is to combine these reactants without losing any solvent and/or reagent and identify the

volatile compound given. In this reaction we specifically used unknow diene # 12 in a eucalyptus

oil sample and the minimum amount of maleic anhydride for this reaction. To determine the

exact amounts used to perform this experiment a gas chromatographic analysis was used to

calculate the percent diene in a 2.5g oil sample, then calculating the minimum amount of maleic

anhydride. The reflux technique was used to prevent loss of solvent during the heating process

and produce maleic anhydride adducts. By performing this technique crystalline products were

produced, then ran through the vacuum filtration technique. The adduct was eventually weighed

and processed through the Digimelt SRS melting point apparatus1 to identify our pure unknown

diene from the oil sample. The experiment yielded 1.65g of product which resulted in a 61.11%

yield recovery. The oil sample #12 melting point range was from 117.2°C- 120°C which is

reasonable to think the sample diene is α-phellandrene.

Introduction

The main focus of this experiment is inspecting the dienes used, specifically a conjugated

diene. A conjugated diene contains double bonds separated by single bonds. The reason why

conjugated dienes are favored is because it allows electron delocalization. This increases stability

for relative dienes like activated alkenes. Conjugated dienes are often found in natural oils and
due to their unique properties of dienes, they provide a strong aroma in the natural oils 3. For

example, eucalyptus oil. It contains various dienes like β-myrcene, allo-ocimene, α-phellandrene

and α-terpinene. In this experiment, the given diene was identified using various techniques.

The Diels-Alder reaction consists of dienes to form Diels-Alder adducts. Particularly

showing a [4+2] cycloaddition product between the alkene and conjugated diene, which has an

electron rich diene and electron poor dienophile (Figure 1). An example that was given is the

Diels-Alder reaction between cyclopentadiene and maleic anhydride to produce a Diels-Alder

adduct1. To further explain this reaction the diene, consists of four pi carbons and the maleic

anhydride consist of two pi carbons to the adducts six-member ring. The reaction is

stereoselective, meaning there are multiple stereoisomers that can form due to the reaction but

prefers the endo formation because it allows more pi electron interaction1.

Figure 1: Diels-Alder reaction with cyclopentadiene (diene) and maleic anhydride (dienophile)

producing a Diels-Alder adduct plus heat

The set up of this experiment starts with calculating the amount of maleic anhydride

needed to add to the oil sample. This was calculated with a given chromatogram analysis. Gas

chromatography is a technique used to analyze liquid mixtures. There are two types of Gas-
liquid chromatography, stationary phase and mobile phase 2. The stationary phase consists of

non- volatile liquid coated on a solid support contained within a column2. The mobile phase is

unreactive carrier gas like He, N2, and Ar etc.2 The way gas chromatography works is by

injecting a liquid sample mixture into the column in a heated injection port. Then the carrier gas

sweeps the components through the column. As the components travel, they are partitioned back

and forth at different rates between liquid and gas phases. As they exit the detector transmits

signal at different rates. The results of the gas chromatography are the different peaks

representing the mixture (Figure 2). The gas chromatograph is sensitive, so it picks up any

substance. For example, there is a particularly small peak shown that isn’t part of the experiment

because it’s an air peak.

Figure 2: Gas chromatography demonstrates various peaks with the largest one being

the main focus

 Given this gas chromatogram of the oil sample, the percent diene in the oil sample can be

calculated. This is done by calculating area for each peak. The percentage is used to figure out

how much diene is in our 2.5g oil sample. Which is then used to calculate the minimum amount

of maleic anhydride needed to allow the reaction to occur.

Afterwards the reflux technique is set up 1. The purpose of this technique is to boil the

substance without losing any solvent. The vapor is then condensed over the hot solution and

returned as liquid in the reaction container. When the reflux is completed, we allow the solution

to cool. Once cooled we collect the adduct and put through the vacuum filtration. This technique

is used to separate liquid-solid mixture by vacuuming all the liquid out of the solid through a

filter paper and drying out the solid. This is done by the vacuum creating a negative pressure in

the filtrate flask which collects the excess liquid. This product is then collected to determine the

identity of the diene in the oil sample. The Digimelt SRS 1 melting point apparatus is used to

determine the melting point of the solid. The experimental melting point was then compared to

the expected melting points to identify the diene in the present original oil sample.

The goal of this experiment is to identify the unknown diene given in the eucalyptus oil by

performing the diel alder reaction through reflux and heat. This technique will allow the solution

to boil then condense and return as vapors until it is ready to then crystalize. It was hypothesized

that our final product was 𝛼-phellandrene because of the purity we concluded due to each

technique.

Experimental Section

To perform this experiment, we need to recognize that the Diels-Alder reactions need to

occur to get the results we need. A gas chromatogram of the unknow oil sample was given by the
lab instructor. The chromatogram demonstrates the largest peak which is the major diene we are

analyzing. By calculating the area of each peak, it was then used to calculate the percent diene.

Which was also used to help calculate how much maleic anhydride was needed for this reaction.

After the amounts were calculated we took 2.5 grams of the unknown oil sample #12, added 5

milliliters of diethyl ether solvent in a round bottom flask. Then we took our calculated 1.13

grams of maleic anhydride and quickly added it to the flask with the other reactants. This step is

proceeded quickly because risking it to moisture in the air can cause our yield and purity of our

product to be compromised. Next, we placed this flask in a hot bath for approximately fifty

minutes with water flowing through the condenser (Figure 3). The condenser is connected at the

top of the flask and the drying tube attached to the condenser, all of which is placed under the

fume hood.

Figure 3: Reflux technique set up with heating used to perform Diels Alder Reactions
 After the time was up, we allowed the contents to cool at room temperature. The contents

slowly crystalized which will indicate transferring to the vacuum filter (Figure 4). We slowly

pour the liquid-solid mixture and wash over the crystals with 10 milliliters of petroleum ether to

help remove the color of the crystals. The solid was now dry and ready to be weighed on the

scale and melted in the Digimelt SRS 1. To identify what the adduct was we compared the

experimental melting point to the theoretical melting points of β-myrcene, allo-ocimene, 𝛼-

terpinene, and 𝛼-phellandrene.

Figure 4: Vacuum filtration set up used to separate liquid-solid mixtures

Results and Discussion:

Before performing all these techniques calculations need to be made to proceed with the

experiment. By using the gas chromatography given, the percent diene in the oil sample needed

to be calculated. This is done by calculating all the peak areas shown. The area is determined by

multiplying the height of each peak and the width of the half height of the peak (Table 1). The

specific equation is:

Peak Area = Height x Width at ½ height

Peak Height (cm) Width at ½ Height Area (cm2)

(cm)

A 1.6 0.5 0.8

B 15.6 0.7 10.9

C 3.3 0.5 1.65

D 1 0.6 0.6

E 5.6 0.6 3.36

Table 1: Measured height, width, and area of five peaks present in the gas chromatography

sample #12

The way each peak was measured was with a small ruler in centimeters. The percent

diene was calculated afterwards by using this specific equation:

𝑎𝑟𝑒𝑎 𝑜𝑓 𝑝𝑒𝑎𝑘
Percent diene = 𝑠𝑢𝑚 𝑜𝑓 𝑎𝑟𝑒𝑎 𝑜𝑓 𝑎𝑙𝑙 𝑝𝑒𝑎𝑘𝑠 𝑥 100

10.92𝑐𝑚2
Percent diene = 0.8𝑐𝑚2 +10.92𝑐𝑚2+1.65𝑐𝑚2+0.6𝑐𝑚2 +3.36𝑐𝑚2 𝑥 100 = 63.01%

Using the percent diene the amount of maleic anhydride can then be calculated. The way this is

calculated is by first figuring out how much diene is in the 2.5-gram oil sample. This is done by

multiplying the percent diene and the mass of samples being 2.5 grams.

Mass of diene = Percent diene x Mass of Sample

Mass of diene = 63.01 x 2.5g= 1.575 grams

The mass of maleic anhydride is then calculated by using a mole-to-mole ratio.

 1 𝑚𝑜𝑙𝑒 𝑢𝑛𝑘𝑛𝑜𝑤𝑛 𝑑𝑖𝑒𝑛𝑒 1 𝑚𝑜𝑙 𝑚𝑎𝑙𝑒𝑖𝑐 𝑎𝑛ℎ𝑦𝑑𝑟𝑖𝑑𝑒 98.06 𝑔
1.575 x 𝑥 𝑥 = 1.13g
136.24 1 𝑚𝑜𝑙 𝑢𝑛𝑘𝑛𝑜𝑤𝑛 𝑑𝑖𝑒𝑛𝑒 1 𝑚𝑜𝑙 𝑚𝑎𝑙𝑒𝑖𝑐 𝑎𝑛ℎ𝑦𝑑𝑟𝑖𝑑𝑒

Following these calculations, we proceeded with the Diels Alder reaction using the reflux

technique and heat. When collecting oil samples and maleic anhydride, a bright yellow appeared

the whole experiment. As the sample cooled at room temperature and crystalized at room

temperature it was ready to filter through the vacuum filtrations. To get rid of the yellow in the

crystals we used 10-15 milliliters of petroleum ether. The final product was weighed and

recorded at 1.65 grams. The theoretical yield was then calculated with the following equation:

𝑚𝑎𝑠𝑠 𝑜𝑓 𝑑𝑖𝑒𝑛𝑒
Theoretical Yield = 𝑥 234.30 𝑔/𝑚𝑜𝑙
136.24 𝑔/𝑚𝑜𝑙

1.575𝑔
Theoretical Yield = 136.24 𝑔/𝑚𝑜𝑙 = 2.7 g Diels Alder Adduct

The percentage yield was then calculated for the experiment

𝑒𝑥𝑝𝑒𝑟𝑖𝑚𝑒𝑛𝑡𝑎𝑙 𝑦𝑖𝑒𝑙𝑑
Percent Yield = x 100
𝑡ℎ𝑒𝑜𝑟𝑒𝑡𝑖𝑐𝑎𝑙 𝑦𝑖𝑒𝑙𝑑

1.65𝑔
Percent Yield = 2.70𝑔 𝑥 100 = 61.11%

The final product was collected and measured in the DigiMelt melting point apparatus. The

melting point range was 119.35°C-122.15°C. We then calculated the average which allowed us to

closely point out the unknow diene being ɑ-phellandrene.

𝑠𝑡𝑎𝑟𝑡 𝑀𝑃+𝑒𝑛𝑑 𝑀𝑃
Average MP = 2

119.35+122.15
Average MP = = 120.75°𝐶
2
Table 2: Diene structures and melting point range for Diel Alder Adducts

The reaction started with 2.5 grams of the unknow oil sample, 1.13 grams of maleic acid and 5

milliliters of anhydrous diethyl ether. It started off very yellow but after the reflux the solution

crystallized and left 1.65 grams of white crystals. There was a theoretical yield of 2.70 grams,

this experiment had a 61.11% yield. Though the crystals were washed with petroleum ether to

clear the color off it didn’t truly need it. The melting point range of the adduct was 119.35°C-

122.15°C. Through all these calculations and measurements, it permitted the experiment to run

smoothly and identify the unknown compound.

Due to there being restrictions during the experiment it caused the adduct to still be

impure. The transfer of the maleic anhydride was quick but because it was so moisture sensitive

it could have encountered moisture in the air and/or the fume hood. If the maleic anhydride came

in contact with moisture can cause yield to be affected negatively and our final adduct to be

inaccurate or out of range for what we are looking for. The reflux technique is meant to be used

with little to no moisture, so it works efficiently. If moisture gets in the way The measurement

of the maleic anhydride was also limited to two decimal places, so the weight of the reactants

wasn’t extremely specific. These disturbances definitely could have impacted on the melting
point of the adduct. Lastly some impurity could have also come from the oil sample because it

doesn’t only contain one diene but other minor dienes in the compound, which may have

produced unintended reactions and different adducts in the final product. But as it was

hypothesized earlier our final adduct did reach the boiling point that matched α-phellandrene,

which demonstrates that the reflux technique, filtration and Digmelt was conducted correctly.

Conclusion:

Ultimately, this experiment produced a Diels Alder Adduct at a melting point of 118.2°C and a

final yield of 61.11%. When compared with different dienes and it’s melting points like β-

myrcene, allo-ocimene, 𝛼-phellandrene, and 𝛼-terpinene. The theoretical melting point being

126-127 degrees Celsius and the experimental melting point is 120.75 degrees Celsius. To get to

this final measurement it started off with the gas chromatography to determine the percent diene

of the unknow oil sample. The diel alder reaction with heating is used to prevent solvent and

reagent loss and finally vacuum filtration to collect the crystal adduct. These techniques

determined the identity of the unknow diene in the oil sample. Due to the disturbances during

these techniques it indicates that the impurity of the sample lowered the melting point. But the

vicinity of the sample melting point demonstrated it being 𝛼-phellandrene the original diene in

the unknow oil sample #12.

Figure 5: The Diels-Alder Adduct of Maleic Anhydride and 𝛼-phellandrene

The objective of this experiment with the following intentions was met by isolating the

compound and creating a low yield of 61.11% due to the reflux technique and vacuum filtration.

The melting point also corresponded with the theoretical melting point of 𝛼-phellandrene

because of the purity of the final adduct.

References:

1. Organic Chemistry Laboratory Manual, Department of Chemistry, Binghamton

University, Binghamton, New York, 2021, pp. 71-74, 110, 116, 134

2. Karugu, R. Organic Chemistry 335 Lecture 3. Binghamton.edu.

https://brightspace.binghamton.edu/d2l/le/content/414394/viewContent/1396178/View

(accessed 2025-03-02).

3. Whitney. Why Is Eucalyptus Oil So Popular And Where Does It Come From? -

AeroWest. AeroWest. https://www.aerowest.com/why-is-eucalyptus-oil-so-popular-and-

where-does-it-come-from/ (accessed 2025-03-03).