Isolation and Purification of Piperine Through Black Pepper

Extraction
Johnathan Harvell
CHEM 440 L01
Undergraduate of Chemistry, Colorado State University, Fort Collins, CO, 80521, United States
09 November 2015
Introduction
Black pepper is a very versatile, and widely used ingredient in culinary arts across the
entire world, and is mainly used for its ability to bring forth a variety of flavors within a food
dish. Scientists have spent many years trying to understand how black pepper has the ability to
create a particular bite in flavor of food from a human perspective, and have found one
molecule being a leading culprit: piperine.1 Piperine is known mainly for its tumor inhibitory
and carcinogenic properties, as well as its inactive nature among other components found in
ground black pepper that exhibit insecticidal properties.2 The synthesis of pure piperine has
been found to be very tedious to perform within the laboratory, and scientists have utilized
certain chemical separation methods to isolate piperine from ground black pepper; however,
the yield of piperine by extraction from black pepper has given reported yields as low as 2-4%.2
The extraction reaction of piperine from natural ground black pepper can be seen
demonstrated below in Figure 1:

Figure 1: Isolation Reaction of Piperine From Natural Ground Black Pepper

The introduction of methylene chloride in the presence of heat with the ground black
pepper allows the piperine to be precipitated into the reaction mixture, and to be separated
from other components found within the black pepper that are may be soluble in hot
methylene chloride. In this experiment, piperine was experimentally isolated from commercial
ground black pepper using the reaction shown in Figure 1 above, checked by TLC, recrystallized,
then purified by flash column chromatography. The isolated piperine product was then
confirmed by 1H NMR and determination of experimental melting point range.
Experimental Procedure3
15 g of ground black pepper was placed in a dry, 100 mL round bottom flask. 30 mL of
CH2Cl2 and five boiling chips were placed inside the flask along with the ground black pepper.
The mixture was refluxed with a water jacketed condenser for 30 minutes. The resulting solid
was extracted from the mixture by vacuum filtration and rinsed with cold CH2Cl2; rotary
evaporation was then used to concentrate the filtrate by removal of any excess solvent. The
solid was cooled in an ice bath, and 10 mL of cold diethyl ether was poured in with the solid,
and mixed together. Rotary evaporation was performed on the mixture for removal of ether
and any excess solvent. The residue was then dissolved in 15 mL of 95% ethanol. The residue
mixture was placed in a 125 mL Erlenmeyer flask with 10 mL of 10% KOH in 95% ethanol. The
solution was heated and water was slowly added to the heated mixture to a total added water
volume of 100 mL using a Pasteur pipette to allow recrystallization. The precipitated, yellow-
brown solid was obtained (crude mass: 0.805 g) from solution by vacuum filtration and washed
with 10 mL of water and 5 mL of cold diethyl ether. The solid was then confirmed for inclusion
of piperine by TLC in comparison to pure piperine in a 40% ethyl acetate/ 60% CH2Cl2 elution
solvent and characterized by UV irradiation of the TLC plate.
The crude piperine product was then purified by the use of the flash column
chromatography using a silica gel base and 50 mL of CH2Cl2 for saturation of the silica gel. The
crude product was dissolved in CH2Cl2 and fractions were obtained with the following v/v ethyl
acetate: CH2Cl2 solvent intervals: 20 mL 10:90, 30 mL 20:80, 30 mL 40:60, and 30 mL 60:40.
Each fraction was investigated using TLC for confirmation of piperine, and the fractions that
were found to contain piperine were added together. The piperine fraction mixture was then
concentrated by rotary evaporation of any remaining solvent. The purified solid (purified mass:
0.895 g) was then characterized by melting point and 1H NMR using a CDCl3 solvent.
Results
In Table 1 below, data regarding the crude and purified yields of the isolated piperine
product from ground black pepper can be seen:
Table 1: Experimental Mass Yield Values of Isolated Piperine

Crude Mass (g) Purified Mass (g) Percent Mass Yield (%)
0.805 0.895 -11.18

The following calculation demonstrates the percent mass yield of the isolated piperine
product by extraction from ground black pepper shown above in Table 1:
0.805 0.895
% = 100% = 100% = 11.18%
0.805

In Table 2 below, the calculated retention factors of the TLC confirmation (as
demonstrated in Appendix A) of the crude, precipitated piperine product before flash column
chromatography can be below:
Table 2: Experimental TLC Retention Factors of Crude Isolated Piperine Product

Spot Solute Distance (cm) Solvent Distance (cm) Retention Factor

Pure Piperine (P) 3.5 5.3 0.66
Co-Spot (C) 3.5 5.3 0.66
Crude Mass (S) 3.5 5.3 0.66
 The following calculation demonstrates the solving of the retention factor for all TLC
spots recorded in Table 2 above:
3.5
= = = 0.66
5.3
In Table 3 below, the calculated retention factors of the TLC confirmation of pure
piperine product within the extracted flash column chromatography fractions (as demonstrated
in Appendix A) can be seen below:
Table 3: Experimental TLC Retention Factors of Purified Piperine Product

SPOT SOLUTE DISTANCE SOLVENT DISTANCE RETENTION FACTOR

(CM) (CM)
PURE PIPERINE (P) 3.0 5.3 0.566
FRACTION # 1 0 5.3 0.000
FRACTION # 2 0 5.3 0.000
FRACTION # 3 4.5 5.3 0.849
FRACTION # 4 4.5 5.3 0.849
FRACTION # 5 3.5 5.3 0.660
FRACTION # 6 3.5 5.3 0.660
FRACTION # 7 3.5 5.3 0.660
FRACTION # 8 3.0 5.3 0.566
FRACTION # 9 3.0 5.3 0.566
FRACTION # 10 3.0 5.3 0.566
FRACTION # 11 3.0 5.3 0.566
FRACTION # 12 2.0 5.3 0.377
FRACTION # 13 1.5 5.3 0.283
FRACTION # 14 1.5 5.3 0.283

In Table 4, the experimental melting point range of purified, isolated piperine product
can be seen compared to the literature melting point range of piperine below3:
Table 4: Experimental Melting Point Range Values of Purified, Isolated Piperine Product

Experimental M.P Range (C) Literature M.P Range (C) Percent Error (%)
103-110 131-135 21.4

The following calculation demonstrates the percent error value of the experimental
melting point range of the purified, isolated piperine product above in Table 4; it is to be noted,
that the percent error value calculated in Table 4 utilizes the largest margin of error in
comparison of the experimental and literature melting point ranges of piperine:
. . 131 103
% = 100 % = 100% = 21.4%
. 131
 In Appendix B, the experimental 1H NMR spectrum of the purified, isolated piperine
product can be seen with the following spectral peaks: 7.419 ppm (s, I = 1, J = 0, F-H), 6.959-
6.964 ppm (d, I = 0.881, J = 1, D-H), 6.854-6.886 ppm (dd, I = 0.416 {6.881-6.886 ppm}, E-H, I =
0.651 {6.854-6.859 ppm}, C-H, J= 4), 6.730-6.770 ppm (t, I = 2.725, J = 2, B-H), 6.404-6.450 ppm
(d, I = 1.013, J = 1, G-H), 5.896-5.956 ppm (q, I = 2.2087, J = 3, A-H), 4.089-4.112 ppm (q, I =
0.219, J = 3, T-H), 3.447-3.616 ppm (q, I = 5.241, J = 3 , J-H), 2.154 ppm (s, I = 0.355, J = 0, Q-H),
1.814 ppm (s, I = 0.616, J = 0 , R-H), 1.569-1.636 ppm (q, I = 7.22, J = 3, K-H), 1.240-1.247 ppm
(d, I = 2.68, J = 1, M-H), 1.168-1.216 ppm (t, I = 0.436, J = 2, X-H), 0.812-0.886 ppm (q, I =2.59, J
= 3 , V-H).
Discussion
As seen in Table 1, it can be found that the purified yield of the isolated piperine
product had a value of -11.18 %, which infers that product after flash column chromatography
was performed still was impure; this is further validated by the NMR spectrum found in
Appendix B, in which the spectral peaks corresponding to Q-H, T-H, V-H, and X-H are in relation
to the presence of hexane and ethyl acetate in the purified product. The presence of ethyl
acetate in the final product would be the result of poor separation during flash column
chromatography of the crude isolated product, while the presence of hexane would be the
result of poor extraction via rotary evaporation during the final phase of purification. It is to be
noted that the product obtained by the combination of flash chromatography fractions after
rotary evaporation was found to be still oily in nature, which could also account of the presence
of ethyl acetate and hexanes found in the experimental NMR spectrum. The presence of ethyl
acetate and hexane in the product would allow a slight increase in overall mass of the purified
product, thus allowing the mass of the purified product being higher in value than that of the
crude product mass found in Table 1. To rid of this experimental error, the utilization of
solvents that do not form an oily composition under the presence of a vacuum would be
recommended for further isolation of piperine, which would allow the solvents to evaporate
and exit the purified mass more successfully. In disregarding the mass that is attributed to the
presence of ethyl acetate and hexanes, the purification yield of the isolated piperine could have
indeed been high in value, which would go against earlier studies showing that the separation
technique found in this study produces low yields of pure piperine of 2-4%.2 To ensure that this
observation is correct, the experiment would need to be replicated multiple times with low
possibility of error resulting in high average purification yield values of isolated piperine from
the same commercial ground black pepper used in this experiment.
During the purification by flash column chromatography, it is to be noted that only
fractions 5-9 were used for determination of melting point, 1H NMR, and purification yield. The
reasoning behind this decision is based on the similarity of the retention factors of fractions 5-9
found in Table 3 in comparison to the retention factors of pure piperine in both Table 2 and
Table 3. It is noticed that the retention factors of pure piperine in both Table 2 and Table 3 are
not equivalent based on measurements made on each TLC plate during the experiment. This
could be the result of an impurity found one of either the TLC plates during the experiment,
which would allow a difference of retention factors to occur. Since the retention factors of
fractions 5-9 range from 0.566-0.66 in correspondence to both retention factors of pure
piperine in Table 2 and Table 3, it was inferred to be the most beneficial choice for the
experiment to combine only these specific fractions; however, due to the probability of error
associated with the TLC of the pure piperine in Table 2 and Table 3, it can be inferred that the
same probability of error can be associated with the combination of the flash column
chromatography fractions 5-9, which could have allowed further impurities to become present
in the purified product. This presence of impurities would then affect the purification yield and
melting point range of the purified isolated piperine found in Table 1 and Table 4. The presence
of additional spots in fractions 5-7 in Appendix A can also be associated with the presence of
impurities in the final product as due to poor separation during flash column chromatography
of the final product.
In Appendix B, it can be seen that alkyl hydrogen at point P could not be identified in the
NMR spectrum due to no present spectral peak found at the expected value of 5.42 ppm.
Assuming that the position of this hydrogen at point P is not equatorial in relation to the
carboxyl group and nitrogen ring in piperine, it can be inferred that the hydrogen is put into a
more axial position that allows a higher probability of collision/attack of that particular
hydrogen in solution, which could demonstrate why it is not found within the NMR spectrum in
Appendix B. Since the purified product was found with traces of ethyl acetate, it can be inferred
that, if ethyl acetate was anionic at some point in solution, then the hydrogen could allow the
quenching of the difference charge; thus, ethyl acetate would be allowed to remain stable in
solution, and piperine would be seen short of this hydrogen in the NMR spectrum as seen in
Appendix B.
The purpose of this experiment was to demonstrate and observe the isolation of
piperine from commercial black pepper by introduction of methylene chloride during reflux,
and purified by the process of recrystallization and flash column chromatography. As seen in
the data obtained in Table 1-4 and Appendices A-B, it can be concluded that isolation of
piperine could possibly result in high purification yield against what has been found in previous
literature and experiments; however, the usefulness of this isolation technique can only be
validated through further replications of this experiment to the point that the probability of
error within the nature of solvents used and other separation techniques performed (such as
rotary evaporation and flash column chromatography) are minimized.
References
(1) Takeda, A.; Tsuboi, S. A New Synthesis of Piperine and Isochavicine. Tetra. Lett. 1979,
12, 1043-1044.
(2) Olsen, R. A; Spessard, G. O. A Short, Stereoselective Synthesis of Piperine and Related
Pepper-Derived Alkaloids. J. Agric. Food Chem. 1981, 29, 942-944.
(3) Somers, P. Experiment #2: Piperine-Isolation and Purification of the Natural Product.
Colorado State University: Canvas.
https://colostate.instructure.com/courses/14977/files/3200968?module_item_id=7614
19 (accessed Oct 2015).
Appendix A: Experimental TLC Plate Series Illustrations

TLC Plate of Pre-Flash Chromatography Crude Material

Flash Column Chromatography of Purification of Isolated Piperine

Appendix B: Experimental NMR Spectrums of Purified, Isolated Piperine
Synthesis and Purification of Piperine by Utilization of Wittig
Reaction Variations
Johnathan Harvell
Undergraduate of Chemistry, Colorado State University, Fort Collins, CO, 80521, United States
CHEM 440 L01
09 November 2015
Introduction
As explored in a previous experiment, piperine is a very versatile substance with many
uses both inside the laboratory and other real world applications, such as its tumor inhibitory
and carcinogenic properties; however, as also demonstrated in previous literature, the isolation
of piperine from ground black pepper has presented low yields from a range of 2-4%.1 Due to
this low yielding of piperine, scientists have spent time creating a series of reactions to allow a
high yield of pure piperine that can be synthesized within a laboratory use low costing starting
materials instead of trying to extracted the molecule from natural products such as raw ground
black pepper. One reaction that has been the most successful and is widely used is the Wittig
reaction, which incorporates a reaction with phosphonium ylides with aldehydes or ketones to
form specific alkenes; Figure 1 below demonstrates the original Wittig reaction mechanism:2

Figure 1: Wittig Reaction Mechanism

The intention of the original Wittig reaction was to create alkenes through the intermediate of a four
membered oxaphosphetane ring, in which the ring would break down due to instability and create the
desired alkene and phosphine oxide.2 The phosphine oxide is created in this reaction due to
phosphoruss affinity for oxygen, which allows resonance of oxygens electrons to form the double bond
in the created alkene.

As time has progressed, the limitations of the Witting reaction have been studied and have been
compensated by the introduction of phosphonate ester carbanions in what is called the Arbuzov
reaction; Figure 2 below demonstrates the mechanism that is associated with the Arbuzov reaction:2

Figure 2: Arbuzov Reaction Mechanism

The reaction utilizes the electrophilic nature of the phosphorous in the trialkylphosphate and the leaving
nature of the alkyl halide to help easily create the necessary phosphorous ester. After the phosphorous
ester is created, a strong base is introduced into the reaction to allow the ester to be deprotonated to
form the carboanion; the carbonanion of the ester is reasonably nucleophilic in solution, which will
allow better probability of the carboanion reacting with introduced aldehydes and ketones in further
steps of synthesis.2 It is to be noticed that minor halide product of the Arbuzov reaction is highly soluble
in water and will allow easier separation of the minor product from the major product during
purification.2

Another further variation of the original Wittig reaction, which also incorporates the use of the
Arbuzov reaction as well, is called the Horner-Wadsworth-Emmons reaction; Figure 3 below
demonstrates the Horner-Wadsworth-Emmons reaction in the presence of an aldehyde:2

Figure 3: Horner-Wadsworth-Emmons Reaction Mechanism

Due to the phosphorous within the carboanion having a high affinity for oxygen, the carboanion
is more likely to attach the carboxyl group of the aldehyde is such a way that a four member
oxaphosphetane ring is formed as seen in Figure 1 for the original Wittig reaction; however,
due to the presence of two oxygen atoms being bonded to phosphorous in the intermediate,
the affinity of phosphorous for both oxygen atoms guide the reaction into producing the trans-
alkene and phosphorous ester as seen in Figure 2.2 The production of the major trans alkene is
based on the stereoselectivity found with the R groups within the four membered ring
intermediate and the preference of the created alkene in being less hindered by allowing the R
groups to be set to opposite sides of the alkene causing a trans-configuration; however, it is to
be noted that cis-configurations of this alkene are still possible, but with very low probability
due to steric hindrance induced by the intermediate in this reaction.2 For the synthesis of
piperine, it is to be noted that the (E,E) stereoisomer is selectively formed due to the same
stereoselectivity demonstrated in the Horner-Wadsworth-Emmons reaction in Figure 2, which
allows black pepper to create the common biting taste that humans experience during
consumption.2
In this experiment, piperine will be synthesized using these variations of the original
Wittig reaction to produce high purified yields of product, and will then be cross-examined and
compared to a previous experiment in the isolation of piperine from commercial black pepper.
The Arbuzov reaction mechanism performed in the synthesis of piperine in this experiment can
be seen demonstrated below in Figure 4, while the Horner-Wadsworth-Emmons reaction
mechanism performed in the synthesis of piperine can be seen in Figure 5:
 Figure 4: Arbuzov Reaction of Experimental Piperine Synthesis

Figure 5: Horner-Wadsworth-Emmons Reaction of Experimental Piperine Synthesis

Experimental Procedure2
Synthesis of Methyl 4-(diethoxyphosphinyl)-2-butenoate
5.3 g (30 mmol, 3.5 mL) of methyl 4-bromo-butenoate was placed in a dry, 50 mL round
bottom flask along with a magnetic stirrer, stirred, and gently heated. 4.8 g (5.0 mL, 30 mmol)
of triethylphosphite is added to the stirred mixture and allowed to undergo a brief exothermic
induction period. After the induction period has ceased, the mixture is attached to a simple
distillation apparatus and temperature is maintained at a constant range of 120-130 C for
approximately one hour or until ethyl bromide is completely distilled from the heated mixture.
Synthesis of Methyl Piperate
0.5 g (22 mmol) of pure sodium metal is placed inside a dry, 50 mL round bottom flask
and allowed to dissolve in 25 mL of absolute methanol to form the necessary sodium
methoxide needed for future steps in the synthesis. During the formation of the sodium
methoxide, the 50 mL round bottom flask is attached to a water jacketed condenser to prevent
the ignition of released hydrogen gas from the reaction. After complete dissolution of the
sodium metal, the reaction flask is allowed to be cooled to room temperature. 5.0 g (4.2 mL, 21
mmol) of synthesized methyl-4-diethoxyphosphinyl-2 butenoate, 3.2 g (22 mmol) of piperonal
and 50 mL of dimethoxyethane were placed inside a dry, 100 mL three-neck round bottom flask
with a magnetic stirrer, thermometer adapter, drying tube and a dropping funnel. The solution
is allowed to be stirred in an ice bath while the created sodium methoxide is added to the
reaction flask via the dropping funnel slowly. After the sodium methoxide is completely in
solution, the flask is warmed to room temperature and stirred continuously for two hours. The
reaction mixture is then poured into 200 mL of cold water and stirred for an additional 45
minutes. The resulting crude solid (crude mass: 1.72 g) is extracted by vacuum filtration and
washed with 25 mL of cold water. The crude product is then recrystallized from ethyl acetate
(purified mass: 1.53 g), and characterized by melting point and 1H NMR.
Synthesis of Piperine
0.2 g (9 mmol) of pure sodium metal is added to a dry, 100 mL round bottom flask with
35 mL of absolute methanol while fitted with a water jacketed condenser to form sodium
methoxide needed for future steps in the synthesis. After the sodium metal is fully dissolved in
the reaction flask, 1.5 g (6.4 mmol) of synthesized methyl piperate and 7.25 mL (72.5 mmol) of
distilled piperidine are added to the reaction flask and allowed to reflux with constant stirring
for 40 hours. The reaction flask is then allowed to be cooled to room temperature. The
resulting red-brown mixture was poured into 100 mL of cold water and stirred for 30 minutes.
The crude solid (crude mass: 2.884 g) is extracted by vacuum filtration and then recrystallized in
an ethyl acetate/hexane solvent mixture. The purified piperine product (purified mass: 1.215 g)
was then characterized by melting point and 1H NMR.
Results
In Table 1 below, the mass yields of synthesized methyl piperate and piperine can be
seen demonstrated below. It is to be noted that the theoretical masses of each synthesized
product is based on the 1:1 molar ratio presented in the reactions found in Figures 4 & 5:
Table 1: Experimental Mass Yield Values of Synthesized Methyl Piperate and Piperine
Synthesized Crude Mass (g) Purified Mass (g) Theoretical Mass Percent Mass
Molecule (g) Yield (%)
Methyl Piperate 1.72 1.53 5.000 30.60
Piperine 2.884 1.215 1.500 81.00

The following calculation demonstrates the percent yield value of the purified piperine
product collected at the end of the experiment as seen above in Table 1:
1.500 1.215
% = (1 ) 100% = (1 ) 100% = 81.00%
1.500
 In Table 2, the experimental melting point ranges of the purified methyl piperate and
piperine are seen compared to literature melting point values below. It is to be noted that the
percent error values in Table 2 are calculated with the highest margin of error associated with
comparison of the experiment and literature melting point range values:
Table 2: Experimental Melting Point Ranges of Purified Methyl Piperate and Piperine
Synthesized Molecule Experimental M.P (C) Literature M.P (C) Percent Error (%)
Methyl Piperate 128-130 145-146 11.7
Piperine 103-105 131-135 22.2

The following calculation demonstrates the calculated percent error value for the
melting point of piperine seen in Table 2 above:
. . 135 105
% = 100 % = 100% = 22.2%
. 135
In Appendix A, the experimental 1H NMR spectrum for the synthesized methyl piperate
and other identified containments contain the following spectral peaks: 7.407-7.442 ppm (d, I =
1, J = 1, Q-H), 7.356-7.391 ppm (d, I = 1, J = 1, G-H), 6.971-6.976 ppm (d, I = 0.907, J = 1, B-H),
6.876-6.908 ppm (dd, I = 0.996, J = 4, D-H), 6.753-6.780 ppm (t, I = 1.57, J = 2, E-H), 6.719 ppm
(s, I = 0.383, J = 0, C-H), 6.668-6.684 ppm (d, I = 0.472, J = 1, F-H), 5.952-5.963 ppm (d, I = 2.35, J
= 1, A-H), 5.902 ppm (s, I = 0.398, J = 0, R-H), 3.745 ppm (s, I = 2.82, J = 0, P-H), 2.154 ppm (s, I =
0.986, J = 0, V-H).
In Appendix B, the experimental 1H NMR spectrum for the synthesized piperine and
other identified containments contain the following spectral peaks: 7.361-7.394 ppm (q, I = 1, J
= 3, Q-H), 7.312-7.346 ppm (q, I = 1, J = 3, E-H), 6.933 ppm (s, I = 0.871, J = 0, J-H), 6.830-6.857
ppm (d, I = 1, J = 1, K-H), 6.684-6.742 ppm (t, L-H {6.684-6.706 ppm}, G-H {6.742 ppm}, I = 2.76, J
= 2), 6.372-6.421 ppm (d, R-H {6.372 ppm}, F-H {6.421 ppm}, I = 1.01, J =1), 4.061-4.085 ppm (d,
I = 0.372, J = 1, D-H), 3.482-3.585 ppm (d, I = 3.81, J = 1, C-H), 2.126 ppm (s, I = 5.33, J = 0, V-H),
1.542-1.610 ppm (dd, X-H {1.596-1.610 ppm}, B-H {1.542-1.555 ppm}, I = 5.82, J = 4), 1.190-
1.214 ppm (d, I = 1.31, J = 1, A-H), 0.787-0.857 (m(5), I = 1.08, J = 3.322, W-H).
Discussion
In Table 1, it can be found that the percent yield of the purified methyl piperate is lower
than the percent yield value of the purified piperine at the end of the synthesis. After
recrystallization of the crude methyl piperate solid, it can be found in Appendix A the possible
inclusion of a carboanion of ethyl acetate corresponding to spectral peak V-H after the
recrystallization of synthesized methyl piperate product. The inclusion of the carboanion could
allow deprotonation of the methyl piperate on the alkyl chain. Due to this deprotonation, it is
possible that the deprotonated products solubility in regards to an aqueous environment could
have been increased, and did not allowed product to be precipitated out during
recrystallization. This would allow a lower yield of purified methyl piperate product as seen in
Table 1. In Appendix A and B, it can be seen that spectral peaks corresponding to Q-H and R-H
are in possible response to the presence of 2(5H)-furanone, which is distinguished by the
manufacturer, Sigma-Aldrich, as a present impurity to approximately 15% of the initial starting
material; however, it is to be noted that the presence of 2(5H)-furanone should have not have
been a factor of error in the performance of these experimental reactions, but could have
contributed to the formation of other unidentified contaminants present in both Appendix A
and B.
In Appendix B, ethyl acetate and hexanes can be identified in the NMR spectrum as
possible contaminants after purification of the crude piperine product. Due to this
contamination, it can be inferred that the high percent yield of purified piperine listed in Table
1 is lower than actually observed. With the inclusion of ethyl acetate and hexane possibly
present in the purified product, the percent error associated with the experimental melting
point range of piperine found Table 2 can also be accounted for as well. This possible
contamination could be due to poor vacuum filtration and rotary evaporation of the purified
product after recrystallization in the ethyl acetate/hexane solvent mixture; however, even with
the exclusion of possible contamination of ethyl acetate and hexane, it can still be reasonably
inferred that the synthesis of the piperine using the Arbuzov and Horner-Wadsworth-Emmons
reactions does present a high yield of purified product, which can be positively compared to
previous literature.2
In comparison to a previous experiment with isolation of piperine from commercial
ground black pepper, it can be seen that percent yield value of the synthesized piperine in Table
1 is much less than that of the percent yield of the isolated piperine (-11.18%); however, as
discussed in the paper regarding the results of the isolated piperine including possible
contamination of the purified product, it can be inferred that the percent yield of the
synthesized piperine in Table 1 is indeed higher and more valid than the reported percent yield
of the isolated piperine from commercial black pepper. The reasonable inference of this
comparison agrees with previous literature that the synthesis route used in this experiment
yields much higher masses of purified piperine than that found in the methods of isolating
piperine used in the previous experiment.12 A potential factor of this high purification yield in
the synthesis route can be attributed to the stereoselectivity found in the Horner-Wadsworth-
Emmons reaction, which allows the major product to be a trans-isomer, thus allowing better
control of the methyl piperate in further steps of the synthesis in response to chemical
environment that it was in. This same stereoselectivity may not have been found in the
isolation of piperine due to the inability to control the stereochemistry of piperine or other
piperine derivatives in response to other molecules that are also found in commercial black
pepper with just the introduction of methylene chloride and heat. As seen in both experiments,
ethyl acetate and hexane were considered to be possible contaminants that may have had an
effect on the precipitation of product during each recrystallization; however, according to the
purification yields of each experiment, the synthetic route seems to have allowed
compensation for the presence of these contaminants, which may allow even more validation
in how synthesis of piperine using the Wittig reaction variations to be more beneficial than the
isolation of piperine from commercial black pepper.
The purpose of this experiment was to demonstrate and observe the synthesis of
purified piperine through the use of variations of the original Wittig reaction and cross-examine
the results against previous experimental data regarding the isolation of piperine from natural
ground black pepper. As discussed in the comparison of both experiments, with the data
presented in Table 1-2 and Appendices A-B, it can be inferred that the synthetic route of
producing piperine results in much higher purification than that found in the previous
experiment of isolating piperine as stated in previous literature; however, further replications
of both experiments would need to be performed with minimized probability to be further
validated.
References
(1) Olsen, R. A; Spessard, G. O. A Short, Stereoselective Synthesis of Piperine and Related
Pepper-Derived Alkaloids. J. Agric. Food Chem. 1981, 29, 942-944.
(2) Somers, P. Experimental # 3: Piperine-Synthesis of a Natural Product. Colorado States
University: Canvas.
https://colostate.instructure.com/courses/14977/files/3200974?module_item_id=7614
20 (accessed Oct 2015).
Appendix A: Experimental NMR Spectrum of Synthesized of Methyl Piperate
Appendix B: Experimental NMR Spectrum of Synthesized, Purified Piperine
Questions
(1) What is the mechanism for the formation of the phosphonate (Arbuzov reaction)?

(2) What is the mechanism for the Horner-Emmons-Wadsworth reaction?

(3) Why is the E-isomer selectively formed?

The E-isomer of piperine is selectively formed due to the stereoselectivity of the R

groups present on the four membered ring intermediate of the Horner-Emmons-
Wadsworth reaction. This stereoselectivity induces the formation of the E-isomer due to
the preferred trans-configuration of the resulting alkene after the release of the
phosphine oxide group.

(4) How would you synthesize the following compounds using a Horner-Emmons reaction?
Show how you would make the required phosphonate. Why would it be better to make
these compounds using the Horner-Emmons Reaction instead of using an aldol-type
reaction?
 An aldol condensation reaction would introduce water into the reaction mixture, which could be
problematic due to the difference in pH of the resulting solution as well as waters ability to
protonate/deprotonate other molecules within the mixture. This may lead to further undesirable
reactions of the created compounds, which may result is an unnecessary product.