Arabian Journal of Chemistry (2016) 9, S537–S540

King Saud University

Arabian Journal of Chemistry

www.ksu.edu.sa
www.sciencedirect.com

ORIGINAL ARTICLE

Double bypasses soxhlet apparatus for extraction

of piperine from Piper nigrum
R. Subramanian a, P. Subbramaniyan b, J. Noorul Ameen a, V. Raj a,*

a
Department of Chemistry, Periyar University, Salem 636 011, Tamil Nadu, India
b
Department of Chemistry, Sona College of Technology, Salem 636 005, Tamil Nadu, India

Received 24 March 2011; accepted 16 June 2011

Available online 25 June 2011

KEYWORDS Abstract A simple modified soxhlet extractor, double bypasses sidearm soxhlet apparatus (DBSA)
Soxhlet; was designed and employed for extraction of piperine from Piper nigrum. Total extraction time,
Modified soxhlet; time taken for a cycle and yield observed in the double bypass sidearm soxhlet apparatus was com-
Piperine; pared with the soxhlet apparatus. Extraction time, time taken for an extraction cycle and yield of
Extraction crude piperine obtained in DBSA were 12 ± 1 h, 8 ± 1.00 min, and 3.90 ± 0.10 g whereas the
results obtained in the soxhlet method were 22 ± 1 h, 16 ± 1.00 min, and 3.80 ± 0.18 g, respec-
tively. The results obtained in DBSA have demonstrated that this approach is as efficient as the
soxhlet apparatus with drastic reduction of extraction time. On the basis of this result, we propose
DBSA as the most efficient method and an alternative to the soxhlet extractor.
ª 2011 Production and hosting by Elsevier B.V. on behalf of King Saud University. This is an open access
article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).

1. Introduction radiotherapy (Sharma et al., 2000). Extraction and preparation

of crude from plant is the starting point for the isolation and
Piperine, one of the major constituents of Piper nigrum purification of chemical constituents (Romanik et al., 2007).
(P. nigrum), has received increasing attention in the recent It is an important step in studies involving the discovery of
years because of its medicinal properties and as bio-availability active compounds of plant materials. Ideally, an extraction
enhancer in formulations of several drugs (Reen and Rashmet, procedure should be exhaustive with respect to the constituents
1997). Piperine shows a protective effect against radiation and, to be analyzed, rapid, simple and inexpensive (Benthin et al.,
therefore, could be administered to cancer patients before 1999).
Soxhlet extraction has been the leaching technique, mostly
* Corresponding author. Tel.: +91 9790694972; fax: +91 427 used for long time extraction and has been a standard tech-
2345124. nique during more than one century and, at present; it is the
E-mail address: alaguraj2@rediffmail.com (V. Raj). main reference to which the performance of other leaching
Peer review under responsibility of King Saud University. methods are compared. Conventional soxhlet apparatus was
originally used for the determination of fat content in milk
(Soxhlet, 1879). In soxhlet extraction, the sample is repeatedly
brought into contact with the fresh portions of the solvent,
Production and hosting by Elsevier thereby helping to displace the transfer equilibrium and no
http://dx.doi.org/10.1016/j.arabjc.2011.06.022
1878-5352 ª 2011 Production and hosting by Elsevier B.V. on behalf of King Saud University.
This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).
S538 R. Subramanian et al.

filtration is required after the leaching step. It is a continuous DBSA. All extractions were continued until colorless. Percent-
process and it required minimum amount of solvent, also the age extraction yield (w/w) for crude was obtained by using the
basic equipment is inexpensive. The most significant demerits formula
of the soxhlet extractor, as compared to the other conventional Weight of crude obtained
techniques for solid sample preparation are, the long time re- Percentage of crude content ¼  100
Weight of sample taken
quired for extraction which causes solvent loss and is harmful
to the environment (Luque de Castro and Garcı́a-Ayuso,
1998). 2.3. Conventional extraction
Several authors have used the soxhlet apparatus for extrac-
tion of pesticides from soil samples and natural products from A weighed sample (40 g) of powdered material was extracted
medicinal plants. Some of the examples for the long extraction at 70–80 C for 24 h under reflux with 1000 mL (250 mL · 4)
process are 16–24, 48–60, 54, and 72 h (Luque-Garcı́a and methanol in a round bottomed flask heated in a water bath.
Luque de Castro, 2003; Chauhan et al., 2004; Brahmachari After extraction the content was concentrated on a water bath
et al., 2006; Amzad Hossain et al., 2006; Perwez and Ali, and the yield of crude was calculated.
2009) depending upon the solid matrixes. The merits and
demerits of the soxhlet apparatus have been used as starting 2.4. Soxhlet and modified soxhlet extractions
point for the development of a variety of modifications to im-
prove the extraction efficiencies. Many attempts have been Exhaustive extraction with methanol (250 mL) was performed
made for the last decades to improve the efficiency of the soxh- in a soxhlet apparatus, the thimble of which contained a
let extractor and bring it closer to that of more recent tech- weighed portion of plant material (40 g). Continuous extrac-
niques such as microwave-assisted solvent extraction tion was performed for about 22 h. The obtained extract was
(Mandal et al., 2009) focused microwave-assisted soxhlet concentrated on a water bath and the yield was calculated. A
extraction (Prados-Rosales et al., 2002) ultrasonic extraction, weighed sample (40 g) of the powdered materials in the thimble
accelerated solvent extraction (Waksmundzka-Hajnos et al., were introduced in to DBSA which was connected with two
2004) high pressure and supercritical fluid extraction solvent distillation flasks through inverted Y shaped joints, as shown
extraction (Adil et al., 2008). in Fig. 3. DBSA extraction Inverted Y-shaped joint was per-
Ultrasound-assisted extraction, shaking extraction and stir- formed with 500 mL of methanol during 12 h.
ring extraction methods are in general less effective than the
soxhlet (Clarke et al., 1991) as they involve most of the soxh- 2.5. Isolation of piperine
let’s disadvantages, but none of its advantages. Some addi-
tional help such as enzymic reaction (Tano-Debrah and The methanol extract on purification over a silica gel (60–120
Ohta, 1995) solvent mixtures have sometimes been coupled mesh) column (60 · 3 cm) using hexane/ethyl acetate step gra-
to the shaking or stirring step in order to improve the overall dients (8:2) afforded crude piperine which was crystallized
efficiency, it hardly surpasses that of soxhlet extraction (Van- using a mixture of 3:2 acetone/hexane and the yield was
Delft et al., 1994). calculated.
The purpose of the present study is to compare soxhlet
apparatus extraction with double bypass sidearm soxhlet
3. Results and discussion
apparatus extraction with respect to extraction time, extraction
cycle, yield of crude piperine and volume of solvent used.
3.1. Optimization of extraction parameters
Methanol is used as the extracting solvent due to its higher sol-
ubilizing capacity. Crude piperine is isolated using column
In this study, the effects of several influential extraction
chromatography.
parameters such as extraction time, volume of solvent and
extraction cycle were systematically studied to set up the opti-
2. Experimental mal extraction conditions to obtain the maximum yield of
crude. Methanol was used as the extracting solvent owing to
2.1. Plant sample its better solubilising capacity for piperine.

Commercial grade sample of P. nigrum was purchased from 3.2. Heat reflux and soxhlet extraction
super market, Yerkaud, Salem, India. P. nigrum fruit berries
were dried and ground to yield powder of 100 mesh size parti- To compare the extraction efficiencies of various modified
cles and were directly subjected to extraction. Methanol, hex- soxhlet methods, soxhlet extraction was used as the reference
ane, petroleum ether, ethyl acetate, diethyl ether and silica method. In heat reflux and soxhlet procedure, extractions were
gel (60–120 mesh for CC and 200–400 mesh for TLC) were continued until the solution becomes colorless (up to 24 h) as
purchased from S.D. Fine Chemicals, Mumbai, India. in the previously reported method (Marion et al., 1966). In
heat reflux, material was extracted with 250 mL of the solvent
2.2. Apparatus for 6 h and the same was repeated four times. In soxhlet
extraction, while heating the flasks, solvent vapors were
Soxhlet (100 mL capacity) was purchased and modified for our brought in contact with the extractant freshly every time,
requirement with the help of a glass fabricator. (Soxhlet single and after leaching, it comes back to the distillation flasks.
bypass: DBSA-double bypasses). Two conventional extraction Frequent addition of fresh solvent increases the solubility
techniques as given below were used for comparison with and leaching. Therefore only one bypass-sidearm exists in
Double bypasses soxhlet apparatus for extraction of piperine from Piper nigrum S539

the soxhlet apparatus (Fig. 1) through which, all vapors have

to go slowly to the extraction tube and hence, extraction is very
slow. About 16 min were required to complete one cycle in
soxhlet extraction, hence for an hour, an extractant was
washed three times by the hot solvent. But in heat reflux meth-
od after 6 h heating followed by filtration, a fresh portion of
the solvent was added until colorless. But in soxhlet extraction
no filtration was required.

3.3. DBSA extraction

An attempt was made to reduce extraction time and increase

the extraction cycle by simple modification in the existing
soxhlet apparatus in such a way that it contains a double by-
pass sidearm (Fig. 2) instead of one bypass sidearm in the Figure 2 Modified soxhlet apparatus (double bypasses).
soxhlet extractor. For the purpose, firstly, DBSA was con-
nected with two distillation flasks via an inverted Y-shaped
joint (Fig. 3). This is called as double bypass soxhlet extractor
with inverted Y-shaped joint (DBSAY). The two sides of the
inverted Y joint were purposefully designed with equal size.
In DBSAY, two distillation flasks were connected at the angle
of 60 slope. Hence there was no horizontal link between two
flasks. When the flasks were heated, double the volume of va-
pors directly reached the extraction tube. Subsequently the
number of extraction cycles per hour was increased and conse-
quently the extraction time was reduced. Traditionally, extrac-
tion of piperine is carried out for extended periods stretching
over 16–24 h (Marion et al., 1966). In DBSAY, the extraction
was completed in 12 h against 24 h in heat reflux and 22 h in
soxhlet extractions. The crude piperine content of the fruits
of P. nigrum is reported to range from 2.8% to 9.0% w/w (Ka-
naki et al., 2008; Anonymous, 1998).

3.4. Comparison of DBSA with other conventional extraction

techniques

The selection of an extraction method mainly depends on the

advantages and disadvantages of the processes, such as extrac-
tion yield, complexity, production cost, environmental friend-
liness and safety. In general, heat reflux extraction is the most
frequently used extraction procedure. The drawback of heat

Figure 3 Double bypasses sidearm soxhlet extractor connected

with two distillation flasks through inverted Y-shaped joint.

reflux extraction is the large amount of solvent, long extraction

times and multistep filtrations needed. Considering the exces-
sive solvent consumption and the long extraction period, this
extraction method is not favorable for commercial perspec-
tives. The drawbacks of soxhlet extraction are long extraction
time and laborious work required. DBSA is a relatively new
method, which has received increasing attention as an alterna-
tive method. DBSA extraction method is the same as conven-
tional soxhlet but with increased extraction cycles with reduced
time. However, compared with the heat reflux and soxhlet
Figure 1 Conventional soxhlet apparatus (single bypass). extraction methods, DBSA method showed prominent
S540 R. Subramanian et al.

Table 1 Extraction time, cycles and yield of crude piperine (n = 3, p < 0.5).
Method of extraction Extraction time (h) Time taken for a cycle (min) Crude piperine yield (%)
SAa 22 ± 1 16 ± 1.00 3.80 ± 0.18
DBSAYb 12 ± 1 8 ± 1.00 3.90 ± 0.10
a
SA: soxhlet apparatus.
b
DBSA: double bypasses sidearm soxhlet with inverted Y-shaped joint.

advantages with high extraction efficiency, reduced extraction Amzad Hossain, M., Islam, A., Jolly, Y.N., Kabir, M.J., 2006. Indian
time which can lead to less laborer work. In the current study, J. Chem. B 45, 1319–1321.
DBSA was compared with the other conventional extraction Anonymous, 1998. The wealth of India, raw materials: Council of
techniques for the extraction of piperine from P. nigrum. On Scientific and Industrial Research, New Delhi.
Benthin, B., Danz, H., Hamburger, M., 1999. J. Chromatogr. A 837,
extraction time, DBSA was also the fastest method with only
211–219.
12 h of extraction time and 24, 22 h in heat reflux and soxhlet
Brahmachari, G., Gorai, G., Chatterjee, D., Mondal, S., Mistri, B.,
extractions. An extraction time, cycle and yield of crude piper- 2006. Indian J. Chem. B 43, 219–222.
ine were given in Table 1. These features along with an ease of Chauhan, S.M.S., Singh, M., Kumar, A., 2004. Indian J. Chem. B 43,
operation and implementation would position DBSA as a sim- 223–226.
ple, fast and economic extraction method with improved effi- Clarke, A.N., Clarke, J.H., Devaney, R.J., DeRoos, F.L., Miille, M.J.,
ciency and reduced extraction time suitable for plant crude 1991. Chemosphere 23, 991–1000.
preparation. Kanaki, N., Dave, M., Padh, H., Rajani, M., 2008. J. Nat. Med. 62,
281–283.
Luque de Castro, M.D., Garcı́a-Ayuso, L.E., 1998. Anal. Chim. Acta
4. Conclusions
369, 1–10.
Luque-Garcı́a, J.L., Luque de Castro, M.D., 2003. J. Chromatogr. A
Modified soxhlet extraction showed that following results: 998, 571–577.
Mandal, V., Dewanjee, S., Mandal, S.C., 2009. Phytochem. Anal. 20,
(i) DBSA is a rapid extraction method for the extraction of 491.
piperine from P. nigrum with clear advantages versus Marion, L., Manske, R.H.F., Holmes, H.L., 1966. The pyrrolidine
heat reflux and conventional soxhlet extraction such as alkaloids. In: The Alkaloids Chemistry and Physiology. Academic
shorter extraction time and lower solvent consumption. Press Inc. Ltd, London.
(ii) The recoveries obtained with the described procedure Perwez, A., Ali, M., 2009. Indian J. Chem. B 48, 443–446.
were almost the same as other methods. Prados-Rosales, R.C., Herrera, M.C., Luque-Garcı́a, J.L., Luque de
(iii) Double bypasses soxhlet apparatus extraction perfor- Castro, M.D., 2002. J. Chromatogr. A 953, 133–140.
Reen, R., Rashmet, K., 1997. J. Ethnopharmacol. 58, 65–173.
mance was satisfactory for P. nigrum extraction hence
Romanik, G., Gilgenast, E., Przyjazny, A., Namiesnik, J., Kaminski,
we propose it is also a suitable and simple method. M., 2007. J. Biochim. Biophys. Meth. 70, 253–261.
Sharma, A., Gautam, S., Jadhav, S.S., 2000. J. Agric. Food Chem. 48,
1340–1344.
Acknowledgment
Soxhlet, F., 1879. Dingler’s Polyt. J. 232, 461.
Tano-Debrah, K., Ohta, Y., 1995. J. Am. Oil’s Chemist Soc. 72, 979–
We are grateful to Mr. C. Janarthanan, Glassware fabricator, 983.
Salem, India for his active support. Van-Delft, R.J., Doveren, A.S., Snijders, A.G., 1994. Fresenius’s J.
Anal. Chem. 350, 638–641.
References Waksmundzka-Hajnos, M., Petruczynik, A., Dragan, A., Wianowska,
D., Dawidowicz, A.L., 2004. Phytochem. Anal. 15, 313.
Adil, I.H., Esra Yener, M., Ayındırlı, A., 2008. Sep. Sci. Technol. 43,
1091.