See discussions, stats, and author profiles for this publication at: https://www.researchgate.

net/publication/301923082

Bromelain: Methods of Extraction, Purification and Therapeutic Applications

Article · May 2016

DOI: 10.1590/1678-4324-2016150010

CITATIONS READS

44 5,256

4 authors, including:

Ali Nawaz Hamid Mukhtar

Government College University, Lahore Government College University, Lahore
50 PUBLICATIONS   155 CITATIONS    175 PUBLICATIONS   1,243 CITATIONS   

SEE PROFILE SEE PROFILE

Ikram ul Haq
Government College University, Lahore
327 PUBLICATIONS   3,797 CITATIONS   

SEE PROFILE

Some of the authors of this publication are also working on these related projects:

PhD project View project

Date fruit Postharvest Management View project

All content following this page was uploaded by Ali Nawaz on 08 June 2016.

The user has requested enhancement of the downloaded file.

 1

Human and Animal Health

Vol.59: e16150010, January-December 2016 BRAZILIAN ARCHIVES OF

http://dx.doi.org/10.1590/1678-4324-2016150010
ISSN 1678-4324 Online Edition BIOLOGY AND TECHNOLOGY
A N I N T E R N A T I O N A L J O U R N A L

Bromelain: Methods of Extraction, Purification and

Therapeutic Applications
Zoya Manzoor1, Ali Nawaz1*, Hamid Mukhtar1, Ikram Haq1.
1
Institute of Industrial Biotechnology GC University Lahore, Pakistan.

ABSTRACT
Bromelain is a concoction of sulfhydryl proteolytic enzymes. Depending upon the site of extraction it can be regarded
as either stem bromelain (SBM) (EC 3.4.22.32) or fruit bromelain (FBM) (EC 3.4.22.33). Bromelain remain enzymatic
active over a broad spectrumand endure a range of pH (5.5 to 8.0) and temperature (35.5 to 71 ºC). It is one of the
extensively investigated proteolytic enzyme owing to its astonishing applications in various industries. This
necessitated employing a strategy that result in highest purified bromelain in less steps and lowest cost. Use of
modernistic approach such as membrane filtration, reverse micellar systems, aqueous two phase extraction and
chromatographic techniques have shown promise in this regard. Besides its industrial applications, bromelain has
been widely utilized as a potential phytomedical compound. Some of its reported actions include inhibition of platelet
aggregation, anti-edematous, anti-thrombotic, anti-inflammatory, modulation of cytokines and immunity, skin
debridement and fibrinolytic activity. It also assist digestion, enhance absorption of other drugs and is a potential
postoperatively agent that promote wound healing and reduce postsurgical discomfort and swelling.

Key words: Phytomedical, Anti-edematous, Fibrinolytic, Anti-thrombotic.

*
Authors for correspondence: [email protected]

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 2 Nawaz, A et al.

INTRODUCTION PROPERTIES

Bromelain is a chief protease enzymes found in Bromelain is actually a group of sulfhydryl

pineapple plant (Ananascomosus) (Smith-Marshall proteolytic enzymes (Smith-Marshall and Golden
and Golden 2012). It has been known chemically 2012) and encompasses variety of cysteine proteases
since 1876 (Tochi et al. 2008) and was identified for (Tochi et al. 2008) when extracted from the stem and
the first time by Marcano in 1891 (Upadhyay et al. fruit of pineapple plant (Neta et al. 2012; Pillai et al.
2010). The investigation and isolation of bromelain 2013). Stem bromelain and fruit bromelain are both
has been started since 1894 (Neta et al. 2012). single-chain glycosylated enzymes but they possess
Sulfhydryl proteolytic enzymes are the chief different characteristics (Barrett et al. 2004). SBM
constituents of bromelain (Tochi et al.2008; Gautam has reduced proteolytic activity and exhibit low
et al. 2010). Bromelain is abundant in stem and fruit specificity for peptide bonds then FBM. Researchers
of pineapple plant and it can also be isolated in small have reported several distinct pH and temperature
amount from pineapple waste such as core, leaves, optima for the activity of bromelain towards its
peel etc. (Hossain et al. 2015). Bromelain present in substrates (De Lencastre et al. 2016). The reported
fruit of pineapple has assigned the EC number EC optimum pH range for SBM by various researchers is
3.4.22.33 and is regarded as fruit bromelain (FBM). 6-7 and its optimum temperature range is 50-60 ºC
Likewise bromelain present in the stem of pineapple (Harrachi et al. 1998; Gautam et al. 2010; Xue et al.
is called stem bromelain (SBM) and its EC number is 2010). While the optimum pH range for FBM is 3-8
EC 3.4.22.32. Stem bromelain possess different and optimum temperature ranges from 37-70 ºC
biochemical properties and composition as compared (Lopes et al. 2009; Jutamongkon and Charoenrein
to fruit bromelain (Pavan et al. 2012) and contains a 2010; Silvestre et al. 2012). Likewise, the molecular
variegated blend of different thiol-endopeptidases. weight range for SBM is 26-37kDa (Grzonka et al.
Efforts are being made by researchers to achieve 2007; Kumar et al. 2011) and for FBM molecular
highly purified bromelain in less steps and low cost. weight range is 24.5-32kDa (Grzonka et al. 2007;
Modern strategies, such as membrane filtration Lopes et al. 2009; Gautam et al. 2010). When
(Lopes et al. 2009), reverse micellar systems bromelain is kept at -20 ºC its stability remains intact
(Upadhyay et al. 2010; Kumar et al. 2011), aqueous for a defined time period (Rowan et al. 1990).
two phase extraction (Coelho et al. 2013; Novaeset Amongst other, cysteine is the most efficient
al. 2013) and chromatographic techniques (Yin et al. compound for the activation of bromelain (Grzonka
2011) have shown promise in this regard. Pineapple et al. 2007). SBM is in fact a combination of several
plant also contains minor quantities of other thiol endopeptidases (Pavan et al. 2012) and also
proteinases like ananain and comosain (Nadzirah et contains compounds like peroxidases, acid
al. 2013) but bromelain is regarded as a primary and phosphatase, several protease inhibitors
extensively investigated component (Amini et al. glucosidases, cellulases, glycoproteins,
2016). The reason of being such valuable is due to its carbohydrates and organically intact Ca2+ (Maurer
miraculous utilization as phytomedical compound 2001; Smith-Marshall and Golden 2012).
(Larocca et al. 2010). Bromelain displays
antiedematous, fibrinolytic, anticancer, anti- EXTRACTION AND PURIFICATION
inflammatory, antibiotic, anticoagulative and METHODS
antithrombotic functions (Kavitha et al. 2013). It is
also a potential postoperatively agent that assists Besides its clinical applications bromelain has been
healing and decrease post surgical discomfort and subjected to many other industries due to its
swelling (Graf 2000). Besides clinical approach, enormous benefits. Researchers thereby are trying
bromelain has also employed in various industries various conventional as well as latest purification
including food industries (Maurer 2001), such as techniques to achieve bromelain in highest purified
breweries (Soares et al. 2012) and flesh processing form at reduced cost (Arshad et al. 2014). As
and tenderization, textile and cosmetic industries compared to fruit, bromelain concentration is high in
(Babu et al. 2008; Ketnawal et al. 2009). With the stem and is thus a cheaply available source of
advent of recombinant DNA technology, scientists bromelain (Tochi et al. 2008). Other parts of
and researchers across the globe have been working pineapple are also investigated for the presence of
on recombinant bromelain to achieve exaggerate and bromelain (Ketnawa et al. 2012) including peel, core
novel applications in future (Arshad et al. 2014).

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 Strategies to obtain bromelain for cure 3

and crown etc. Extraction of bromelain from these exposed to numerous purification stages to eradicate
parts is attractive not only from environmental point impurities that may interfere with bromelain to
of view but also economically (Novaes et al. 2013). hinder its application and reduce the specific activity
Bromelain can be easily extracted from the juice of of the enzyme (Illanes 2008). Product purity is the
pineapple by ultrafiltration (Larocca et al. 2010) but key factor which may constitute a large proportion of
still FBM is not commercially available due to being the total enzyme production cost (Lightfoot 1990).
different from SBM (Pavan et al. 2012). The Several conventional isolation and purification
marketable bromelain is mostly extracted from the techniques are now obsoleted because of their low
stem of pineapple through centrifugation, purification potential (Soares et al. 2012).So the
ultrafiltration, lyophilization (Corzo et al. 2011) and extraction and purification strategy (Figure 1)
two-step Fast Protein Liquid Chromatography designed should be selective for the desired product,
(FPLC) (Harrach et al. 1998). Once extracted, the cheap, high yielding and speedy (Gupta et al. 2004).
crude mixture containing required enzyme is then

Figure 1 - Overview of extraction and purification of Bromelain.

Due to increased interest of scientists toward 5) Different chromatographic techniques

bromelain, several new purification techniques have (Devakate et al. 2009; Li et al. 2009; Paulo et al.
been employed for its extraction and purification 2012).
(Table no. 1). These include:
1) Aqueous two phase systems (Babu et al. Reverse micellar system
2008; Ferreira et al. 2011; Coelho et al. 2013; Spir et Micelle is an aggregate of molecules possessing both
al. 2015). polar and non-polar regions. Reverse micelles are
2) Membrane processes (Doko et al. 2005; thermodynamically stable, minute surfactants that
Lopes et al. 2009) hold water in their interior surrounded by organic
3) Precipitation (Doko et al. 2005; Devakate phase (Andray et al. 2001). Only protein of interest is
et al. 2009; Soares et al. 2012). entrapped in micelle (Figure 2) whereas other
4) Reversed micellar systems (Hemavathi et impurities remain in organic phase (Lee and Chong
al. 2007; Navapara et al. 2008; Kumar et al. 2011). 2011). Reverse micellar system is an encouraging
strategy for downstream processing. It is ideal to

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 4 Nawaz, A et al.

extract biomolecules through diluted samples. with ultrafiltration was also studied which resulted in
Reverse micellar system possess higher sample an activity recovery of 95.8 % and purification factor
loading capacity, are specific and easy to operate after ultrafiltration of 8.9-fold (Hebbar et al. 2012).
(Chen et al. 2008). The use of affinity based reverse micellar extraction
RMS are also being used for the extraction and and separation technique to extract and purify
purification of bromelain from ananas stem and bromelain from pineapple waste yielded purification
waste. Pineapple core yielded 106% active recovery of 12.32 fold with an activity recovery of 185.6 %
and 5.2 purification fold (Hebbar et al. 2008). There (Kumar et al. 2011). Moreover the process
is also reported use of RMS for extraction of optimization studies for RMS had been reported
bromelain from pineapple where it yielded 97.56% (Fileti et al. 2009; Dhaneshwar 2014) and both the
activity recovery and 4.54 purification fold batch and continuous extraction of bromelain from
(Hemavathi et al. 2011). Researchers are pineapple juice by under optimized conditions was
implementing various modifications in RMS to also studied (Fileti et al. 2007). A neutral model
upgrade the yield and purification fold. To purify developed for bromelain extraction from pineapple
bromelain at larger scale, scale-up studies by phase juice by RMS under optimized conditions gave 4.96
transfer mode of reverse micellar system are also purification factor with productivity of 1.29 ml/min
performed which yielded purification fold of 2.43 (Fileti et al. 2009).
with an activity recovery of 81.3% (Hebbar et al.
2011). To uplift the efficacy of process RMS coupled

Figure 2 - Stages in working of Reverse Micellar System (RMS).

Aqueous two phase system (ATPS) used in initial stages (Gupta et al. 1999). High active
The aqueous two-phase systems consists of a recovery of enzymes during ATPS is due to the
polymer and a salt (or two polymers) and have been presence of PEG which causes alteration in the
regarded as extensively used tools for extraction and structure of active sites of the enzyme. Researchers
purification of biomolecules (Rabelo et al. 2004). It have reported the used ATPS for extraction and
is low cost, rapid, possess reusable polymers purification of bromelain from stem and peel of
(Johansson et al. 2011), scalable (Asenjo and pineapple (Ferreira et al. 2014).
Andrews 2011) and can withstand high biomass load Extraction and purification of bromelain complexed
as compared to rest of the separation systems (Kaul with polyphenol oxidase through ATPS from the
2001). ATPS is reported as a challenging technique pineapple resulted in 228% active recovery with 4
for the separation of molecules that are hard to be fold rise in purification (Babu et al. 2008). ATPS has
separated through other methods and preferred to be been used to recover bromelain from the peel of

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 Strategies to obtain bromelain for cure 5

pineapple and gave 113.54% and 206% active equilibrium and ATPS were employed together for
recovery with 2.23 and 3.44 purification fold the purification of bromelain, purification fold came
(Ketnawa et al. 2010). In order to optimize the out to be among 25-62 (Ferreira et al. 2011).
separation of bromelain response surface Moreover, thermoseparation of bromelain resulted in
methodology has been worked in ATPS that yielded active recovery of 79.5% with 1.25 purification fold
90.33% enzyme and its purification factor was 2.4 (Rabelo et al. 2004).
(Navapara et al. 2011). When thermodynamic

Figure 3 - Extraction and Purification of Bromelain through Aqueous Two Phase System (ATPS).

Chromatographic techniques (Devakate et al. 2009). High-speed counter-current

Chromatographic techniques significantly conserve chromatography (HSCCC) has also been used to
the structure of purified protein (Gautam et al. 2010). purify bromelain which generated 3.01g bromelain
Several chromatographic techniques such as ion from 5.00g crude extract (Yin et al. 2011).
exchange chromatography (Hung et al. 2002; Immobilized metal affinity membrane (IMAM) has
Arumugam and Ponnusami 2013; Iara et al. 2013; been employed to purify bromelain which resulted in
Swaroop and Viswanathan 2013), high-speed 15.4 fold purification factor with 94.6% recovery
counter-current chromatography (HSCCC) (Yin et (Nie et al. 2008). 87.4% active recovery of bromelain
al. 2011), affinity chromatography, gel filtration was achieved when Poly Acryl Acid (PAA)-bound
chromatography and capillary iron oxide magnetic nanoparticles were used in order
electrochromatography (Cheng and Huang 2004; to adsorb bromelain from aqueous solution (Cheng
Babu et al. 2008; Chen et al. 2008) have been and Huang 2004). In the same context, 13 fold
employed for the purification of bromelain. All these purification factor was achieved when bromelain was
techniques possess different specifications thus result adsorbed in expanded bed (Silveira et al. 2009). 3.3
in different purification efficiency and recovery (Yin fold enhanced purification of bromelain was
et al. 2011). Amongst these, ion exchange achieved by the combinations of precipitation and
chromatography (IEX) is extensivelyemployed for chromatographic techniques (Devakate et al. 2009).
bromelain separation (Devi and Sowmiya, 2014). Ion
exchange chromatography is highly specific, Purification through membrane filtration
scalable, reliable and a cheap purification method Membrane filtration employs the use of membranes
(Ng et al. 2009). 10 fold purification of bromelain to purify molecules on the basis of size difference. It
was achieved using cation exchange chromatography includes microfiltration and ultrafiltration and is

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 6 Nawaz, A et al.

considered quite useful for enzymes seclusion as well recovery and purification fold. Ammonium sulfate
as concentration (Cassano et al. 2003). Membrane precipitation is a cheap and extensively used method
filtration is being used for the isolation and for protein purification (Saxena et al. 2007).
purification of bromelain. Simultaneous use of Ammonium sulfate precipitation resulted in 2.81
microfiltration and ultrafiltration in order to separate purification fold of bromelain (Devakate et al. 2009).
bromelain from pineapple pulp resulted in active In a study ethanolic extraction, isoelectric focusing
recovery of 85% (through microfiltration) and 10 and ammonium sulfate precipitation were compared
fold concentrated bromelain (through ultrafiltration) for the extraction of bromelain. Amongst these three,
(Lopes et al. 2009). When microfiltration, ethanolic extract resulted in highest bromelain yield
ultrafiltration, ammonium sulfate precipitation and (Silvestre et al. 2011). Likewise, a comparison was
ultracentrifugation were employed in a sequence also made between ethanol, PEG and (NH₄)₂SO₄ to
98% yield was achieved (Doka et al. 2005). precipitate bromelain in order to recover it from stem,
Membrane filtration in conjugation with other bark and leaves of pineapple plant. The purification
processes results in enhanced purification of factor achieved after using ethanol was 2.07 fold with
bromelain. When stem bromelain was adsorbed on 30-70% recovery. 20-40% recovery and 4.4 fold
nano-TiO2 and then passes through ultrafilters, 5.3 purification factor was achieved after precipitation by
purification fold and 64.7% active recovery was ammonium sulfate whereas poly (ethylene glycol)
achieved (Chao et al. 2009). In a study three methods failed to precipitate bromelain (Soares et al. 2012).
(Kaolin adsorption, ultrafiltration and tannin 1400 GDU/ml proteolytic activity with a protein
precipitation) were performed and compared for content of 9.33 mg/ml was achieved after using
bromelain purification. Bromelain with highest homogenization process for bromelain separation
proteolytic activity was obtained by ultrafiltration (Li (Loh et al. 2005). The use of recyclable mesoporous
et al. 2009). silica to adsorb bromelain from pineapple juice
resulted in 6.2 fold purification with 97.89%
Other methods recovery (Arumugam and Ponnusami 2013).
Various other methods have also been worked to
separate bromelain and resulted in enhances active

Table 1 - Some modern techniques for the extraction and purification of Bromelain

Purification Types Yield Purificat Reference

techniques (%) ion
fold/fact
or
PEG/K2So4 aqueous two-phase 228 4.0 Babu et al. 2008
system
PEG/poly(acrylic acid) aqueous 335.2 25.78 Novaes et al. 2013
two-phase system 7
PEG/(NH₄)₂SO₄ unconventional - 11.80 Coelho et al. 2013
aqueous two-phase system
PEG/MgSo4 aqueous two phase 113.5 2.23 Ketnawa et al. 2010
Aqueous two- system 4
phase system PEG/MgSo4 aqueous two phase 108.4 - Katnawa et al. 2009
system 5
PEG/MgSo4 aqueous two phase 206 3.44 Ketnawa et al.
system 2011a
Block copolymers aqueous two 79.5 1.25 Rabelo et al. 2004
phase system
PEG/MgSo4 aqueous two phase - 25-62 Ferreira et al. 2011
system
Four various aqueous two phase - - Ketnawa et al.
system 2011b

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 Strategies to obtain bromelain for cure 7

Microfiltration & ultrafiltration 85- 10 Lopes et al. 2009

Filtration 100
Microfiltration, ammonium sulfate 50 - Doka et al. 2005
precipitation, ultracentrifugation
Nano-TiO 2, ultrafiltration 64.75 5.3 Chao et al. 2009
Reverse micelle systems 85 4 Chaurasiya et al.
2015
Reverse micelle systems 102 5.2 Hebbar et al. 2008
Reverse Reverse micelle systems 97.56 4.54 Hemavathi et al.
micellar 2007
systems Optimized reverse micelle systems 89.6 2.8 Navapara et al.
2011
Affinity-based reverse micelle 185.6 12.32 Kumar et al. 2011
system
High speed counter-current - - Yin et al. 2011
chromatography, reverse
micelle system
Chromatogra Immobilized metal affinity 94.6 15.4 Nie et al. 2008
phy membrane
Precipitation, ion exchange - 3.3 Devakate et al.
chromatography 2009
Cation exchange chromatography - 10 Nadzirah et al.
2013

APPLICATIONS OF BROMELAIN of which the tissues appear to be inflated and

Bromelain has been known for its vast commercial inflamed (Jaber 2002). Mostly NSAIDS (non-
applications. It is being used in food and beverage steroidal anti-inflammatory drugs) are prescribed to
industries (Neta et al. 2012), in meat tenderization, in
ward off the classical signs of inflammation (Calor,
cosmetic industries (Orsini 2006) as well as in textiledolor, rubor and tumor) (Charles et al. 2011),
industries (Arshad et al. 2014). Besides its industrialhowever these drugs severely injures GIT
applications, bromelain possess multiple therapeutic (Gastrointestinal tract) and possess numerous
actions (Summarized in Table no. 2). Some of its hazardous after effects (Walker et al. 2002) so, as an
reported actions include inhibition of platelet alternative bromelain has been demonstrated as an
aggregation, anti-edema (MacKay et al. 2003), anti- anti-inflammatory agent and is in practice for acute
thrombotic and fibrinolytic activity (Errasti et al. inflammation and several related conditions (Hale et
2016), anti-inflammatory action (Brien et al. 2004), al. 2005a). Anti-inflammatory action of bromelain is
anti-tumor action (Dhandayuthapani et al. 2012; mediated by retarding the formation of pro-
Pillai et al. 2013), modulation of cytokines and inflammatory prostaglandins (Graf 2000), reducing
immunity, skin debridement properties (Hu et al. the cell surface receptors such as hyaluronan receptor
2011; Rosenberg et al. 2012), enhanced absorption of CD44 and plasma fibrinogen levels (Yuan et al.
other drugs (Orsini 2006), mucolytic properties, 2006). It also reduces the level of bradykinin, CD4+
digestive assistance, enhanced wound healing T lymphocytes (Manhart et al. 2002) and enhances
(Taussig et al. 1988), cardiovascular and circulatory the activity of serum fibrinolytic thus induces the
improvement (Kavitha et al. 2013). Some of its production of proinflammatory mediators and anti-
therapeutic applications are briefly discussed below. inflammatory prostaglandins as a result of which the
distention and permeability of capillaries is reduced
Anti-inflammatory agent (Darshan and Doreswamy 2004; Hale et al. 2005;
Inflammation is the body's attempt of self- Tochi et al. 2008). Figure no. 4 displays how
protection which takes into account the discharge bromelain respond to certain mediators of acute
of various factors that causes the distention of inflammation.
capillary and enhances their permeability, as a result

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 8 Srivastava, AK et al.

(IL-1)
Leukocytes
adhession
(PGI2)
(TNF)
Vasodilation
, Inhibit Leukocytes
platelets adhession
aggregation

Bromelain

(PGE2)
(Bradykinin)
Vasodilation
Vascular
permeability, (Thrombox
pain ane A2)
Platelets
aggregation,
vasoconstrict
ion

Figure 4: Effect of Bromelain on certain mediators of acute inflammation

Rhinitis and rhinosinusitis 2004). Recently it has been found that bromelain,
Rhinosinusitis results in inflammation of nasal cavity when used in conjugation with other nutraceuticals
and paranasal sinuses (Guo et al. 2006) whereas such as turmeric, results in enhanced efficacy in the
chronic rhinosinusitis (CRS) is more persistent and treatment of degenerative joint pain diseases
long-term inflammation which causes disruption of (Conrozier et al. 2014).
mucus membranes (Mehdi et al. 2014). Bromelain is Colonic inflammation
an effective mucolytic agent and is being efficiently Bromelain notably reduce the harshness of colonic
used in rhinitis, rhinosinusitis as well as in chronic inflammation and when taken orally, it significantly
rhinosinusitis (Guo et al. 2006; Buttner et al. 2013). reduces the severity of ulcerative colitis (Hale et al.
It decreases the formation of pro-inflammatory 2005). The chief anti-inflammatory mechanism of
prostaglandin and reduces swelling in nasal passages bromelain comes out to be proteolytic in nature by
(Helms and Miller 2006). Bromelain also decrease which it eradicates cell surface receptors involved in
mucus production and aids its drainage (Tochi et al. leukocyte defection and activation (Hale et al. 2010).
2008). Beneficial effects of bromelain have been Bromelain also vary the emission of certain
viewed in allergic rhinitis as well where it reduces chemokines and thus reduces the prevalence and
edema and inflammation (Thor and Kelly 2000). sternness of spontaneous colitis. Due to these
researchers regard bromelain as a novel treatment for
Arthritis inflammatory bowel disease (Onkan et al. 2008).
NSAIDS are commonly prescribed in arthritis
(Henriksson et al. 2014). Bromelain can be used as Anti-cancer agent
harmless alternative and researchers have found that In healthy cells, autophagy occurs naturally whereas
bromelain exhibits promising efficacy in arthritis in tumor cells, this process is deactivated. Cancerous
(Osteoarthritis and rheumatoid arthritis) (Pavan et al. cells metastasize through circulation and transport
2012). Osteoarthritis, the major form of arthritis, is a system to nearby tissues. In many types of cancer p53
leading cause of disability these days. Bromelain is a gene, which induces apoptosis, is inactivated thus
potent analgesic and displays a direct effect on apoptotic cell death is evaded in such cancerous cells
certain mediators of pain like bradykinin (Brien et al. (Baez et al. 2007). Researchers have recognized

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 Strategies to obtain bromelain for cure 9

numerous anti-cancer mechanisms of bromelain such Akt (a protein kinase) controls numerous cellular
as switching off the essential gene signal NF-kappaB, processes and is a key player in cardiovascular
suppression of Cox-2 expression, upregulation of p53 diseases. It is activated upon phosphorylation
and Bax and by initiation of autophagy (Tochi et al. (Chaanine and Hajjar 2011). Apoptosis is a chief
2008; Bhui et al. 2010). Bromelain exhibits stimulator of several cardiovascular disorders
antimetastatic activity, stimulates several caspases (Maulik and Das 1994). Upon its activation, Akt
and promotes apoptosis. Moreover the use of inactivates several proapoptotic genes that causes
bromelain in cancer lessens tumor size and results in cessation of apoptosis to some extent resulting in
reduced damage to healthy cells and possesses less cardioprotection (Shiraishi et al. 2004). Bromelain is
after effects as compared to chemotherapy (Bhui et an effective cardioprotective agent. It possesses
al. 2009; Amini et al. 2016). Bromelain remarkably antithrombotic and anticoagulant activities and
results in tumor regression for certain cell lines. reduces platelet aggregation. It also prohibits the
These include P-388 leukemia, sarcoma (S-37), A- attachment of platelets to endothelial (Metzig et al.
431 epidermoid carcinoma, A-375 melanoma, lewis 1999). Bromelain results in enhanced
lung cancer (Batkin et al., 1988) and ADC-755 breast phosphorylation of Akt which causes inhibition of
cancer (Bhui et al. 2011; Pavan et al. 2012). apoptotic cell death (Juhasz et al. 2008). Bromelain
Furthermore, the use of bromelain in conjugation inhibits clumping of platelets and also put a stop to
with chemotherapeutic medicine improves the thrombus formation (Metzag et al. 1999; Bhatacharia
efficiency of these drugs to certain extent (Pillai et al. 2008). It has the ability to reduce angina and also
2013; Amini et al. 2014). possess antihypertensive action (Maurer 2001).

Breast Cancer Angina Pectoris and Thrombophlebitis

Bromelain, a pharmacologically active compound, is Bromelain has been effective in the treatment of
a potent anti-tumorigenic agent (Neta et al. 2012). In cardio vascular diseases (Tochi et al.2008).
mammary carcinoma cells bromelain affects MCF-7 Bromelain promotes the disruption of thrombus,
cells by slowing down their growth inhibitory reduces the platelets clumping, blood viscosity and
response and activate the process of autophagy. thus reduces the harshness of angina pectoris (Metzik
Furthermore, bromelain promotes natural cell death et al. 1999;Maurier 2001) and transient ischemic
(apoptosis) in cancerous cells (Bhui et al. 2010). attack (TIA). Bromelain also helps to reduce the
When taken orally, it also encourages the scarce potent risks of thrombophlebitis and aids its
monocytic cytotoxicity in breast cancer patients treatment (Pavan et al. 2012). Bromelain is also
(Eckert et al. 1999). Mainly in mammary cancer cells recommended for the treatment of acute
increased dosage of bromelain promotes the process thrombophlebitis (Ley et al. 2011).
of natural cell death (apoptosis) (Dhandayuthapani et
al. 2012). Burns Debriding agent
A dry Scab formed on the skin especially after burn
Epidermoid carcinoma and Melanoma is known as eschar which may result in prolonged
The efficacy of bromelain against particular recovery and also enhance the chance to develop
melanoma and epidermoid carcinoma cell lines was infection (Singer et al. 2010b). Debridement refers to
checked where bromelain successfully exhibited the elimination of dead, damaged, or infected tissue
anti-cancer activity. Bromealin not only minimized from the site of eschar to improve the healing
their proliferation and reduced the expression of Cox- potential of the remaining healthy tissues (Rosenberg
2 gene (Bhui et al. 2012) but also induced apoptosis et al. 2012). Debriding the burn eschar surgically is
and suppressed lung metastasis of melanoma cells quite challenging and can cause many difficulties
(Carla et al. 2007). CD44 are the surface proteins (Pavan et al. 2012). On the other hand; enzymatic
present on human leukemia Molt 4/8 cells and are debridement can minimize such complications and is
involved in cancer matastasis. Bromelain efficiently a safe alternative (Krieger et al. 2012). Amongst
reduced the volume of this protein in leukemia and these, bromelain is regarded as a potential candidate
melanoma cells (Harrach et al. 1994). (Ahle and Hamlet 1987; Maurer 2001). Topical
bromelain is being used these days in debriding
Inhibitor of thrombus formation necrotic tissues (Hu et al. 2011; Wu et al. 2012).
Platelets aggression and thrombus formation are the Bromelain encompasses a fraction of Escharase
leading factors of cardiac discomfort (Bousser 2013). which is a main debriding agent (Pavan et al. 2012).

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 10 Nawaz, A et al.

Bromelain is quite specific in its action. It selectively as changes the proportion of CD4+/CD8+ (Secor et
acts upon the effected tissues without harming the al. 2005a; 2007b; 2012c; 2013d).
normal ones (Rosenberg et al. 2012). Bomelain
simplifies the debridement process and offers Blood Coagulation and Fibrinolysis
improved and accelerated healing and quick Fibrinolysis is a process to prevent fibrin clot from
reepithelialization (Singer et al. 2010a; Hu et al. growing and allows the body to clear fragments of
2011; Rosenberg et al. 2012).Researchers have found clots safely (Maus and Hajjar 2005). Bromelain is
bromelain to be useful in healing post-surgical regarded as an effective fibrinolytic agent and
wounds and aid to lessen their aching and swelling prevents blood from coagulation (Taussig and Batkin
etc (Graf 2000; MacKayand Miller2003). 1988) by exaggerating the transformation of
plasminogen to plasma which in turn inhibit fibrin (a
Asthma protein involved in blood clotting) synthesis
Asthma refers to the inflammation of air passages (Bhattacharia 2008). It also decreases the proportion
resulting in swollen or inflamed airways. of fibrinogen in serum,represses ADP induced
Inflammation in asthma is caused mainly due to platelet aggregation and delay both prothrombin time
amplification of eosinophils and T lymphocytes level (PT) and activated partial thromboplastin time
in the bronchial mucosa and broncho-alveolar lavage (APTT) (Kaur et al. 2014; Errasti et al. 2016). Both
(BAL) fluid (Leblond et al. 2005). Bromelain is an extrinsic and intrinsic pathway results in the
efficient therapeutic agent and is being used formation of fibrin ( as shown in Figure no. 5).
successfully in allergic airway disease (AAD) (Secor However bromelain limits its formation by reducing
et al. 2012). Bromelain treats asthma by decreasing some of the intermediates of clotting cascades (factor
the level of total BAL leukocytes (eosinophils and X and prothrombin) and increase fibrinolysis. It also
lymphocytes) and cellular infiltrates (Secor et al. decreases prekallikrein and thus inhibits the
2005) and also notably lessens BAL CD4+, CD8+ T generation of bradykinin at the site of inflammation.
CD4+ and CD25+T cells (Jaber 2002; Darshan and This results in reduction of pain and edema as well as
Doreswamy 2004). It also reduces interleukins IL-4, increases circulation at the site of injury.
IL-12, IL-13, IL-17 and IFN-α in the serum as well

Figure 5: Pathways through which Bromelain inhibits blood clotting.

Dermatological disorders After treatment all the victims of PLC were fully
Pityriasislichenoideschronica (PLC) recovered with zero side effects which is a virtue of
Pityriasislichenoides is a rare cutaneous disorder and its immuno-modulatory and antiviral trait
is characterized by the development of multiple, (Massimiliano et al. 2007).
scaly, erythematous to brown papules on the trunk
and extremities (Someshwar and Udare 2012). Scleroderma
Bromelain has been investigated for use in PLC.

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 Strategies to obtain bromelain for cure 11

It is a disease characterized by progressive skin Bromelain has also shown promise in treating
hardening and induration that is caused by abnormal scleroderma (Gaby 2006).
growth of connective tissues (Gabrielli et al. 2009).
Table no. 2: Summary of clinical applications of Bromelain.
Disorders Observed effects References
Asthma Change CD4+ to CD8+ T Jaber et al. 2002;
lymphocyte ratio and decreases AAD Secor et al. 2008
(Allergic airway disease).
Chronic Retards formation of pro- Bakhshaee et al.
Anti-Inflammatory Rhinosinusitis inflammatory prostaglandin resulting 2014
agent in fast recovery.
Colonic Decreased the occurrence and Darshan;
inflammation severity of spontaneous colitis Doreswamy 2004
Osteoarthritis Reduces in joint tenderness, pain and Klein 2006; Walker
swelling et al. 2002
Rheumatoid Reduce joint stiffness Maurer 2001
arthritis
Soft tissue injuries Have high wound healing capacity Baumann et al.
2007; Lemay et al.
2004
Breast cancer Decrease tumor size and cause Baez et al. 2007
apoptosis
Leukemia Causes tumor regression Maurer 2001;Pavan
et al. 2012
Anti-tumor agent Lung carcinoma Bromelain possesses antimetastatic QIMR 2005
and anticoagulant functions
Ovarian cancer Decrease tumor growth and invasive Maurer 2001
potential
Melanoma Inhibits nuclear factor-kappab (NF- Kalra et al. 2007
kb), a protein involved in cancer
Postoperative Recovers abdominal distention and Wen et al. 2006
Improvement of gastrointestinal increase water content of fecal pellets
Gastrointestinal dysmotility (ileus)
Tract related Constipation Improves stool release in patients Wen et al.2006
discomforts Exocrine pancreas Improves digestion Knill-Jones et al.
insufficiency 1970
Nausea, vomiting, Neutralizes the effects of Pavan et al. 2012
diarrhea intestinal pathogens that causes NVD
Angina pectoris Bromelain stops aggregation of Taussig and Nieper
Inhibition of platelets and causes blood thinning 1979
thrombus Transient ischemic Reduces its severity Taussig and Nieper
formation attacks 1979
Thrombophlebitis Treats thrombophlebitis Kelly 1996
Thrombosis Break down cholesterol plaques Kelly 1996
Burns/Eschar Topical bromelain results in Rosenberg et
Treatment of complete debridement al.2004
Dermatological Wrinkles Conditions skin Reddy et al. 2013
disorders Pityriasislichenoide Complete clinical recovery Massimiliano et al.
schronica 2007
Scleroderma Resolves lesions Gaby 2006

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 12 Srivastava, AK et al.

CONCLUSION Barrett AJ, Rawlings ND, Woessner JF. Handbook of

Proteolytic Enzymes. 2. ed. Amsterdam: Elsevier
Bromelain is a concoction of several thiol Academic Press, 2004. p. 1051-1071.
Batkin S, Taussig SJ, Szekerezes J. Antimetastatic effect
endopeptidases. It is distinguished as stem bromelain
of bromelain with or without its proteolytic and
and fruit bromelain depending upon the site of anticoagulant activity. J Cancer Res Clin.1988; 114(5):
extraction. It is one of the extensively investigated 507-508.
proteolytic enzyme owing to its astonishing Baumann LS. Botanical ingredients in cosmeceuticals. J
applications in various industries. This necessitates to Drugs Dermatol. 2007; 6(11): 1084-1088.
employ a strategy that result in highest purified Bhui K, Prasad S, George J, Shukla Y. Bromelain inhibits
bromelain in less steps and lowest cost. Use of COX-2 expression by blocking the activation of
modernistic approach such as membrane filtration, MAPK regulated NF-kappa B against skin tumor-
reverse micellar systems, aqueous two phase initiation triggering mitochondrial death pathway.
extraction and chromatographic techniques have Cancer Lett. 2009; 282(1): 167-176.
Bhui K, Tyagi S, Prakash B, Shukla Y. Pineapple
shown promise in this regard.
bromelain induces autophagy, facilitating apoptotic
response in mammary carcinoma cells. Biofactors.
REFERENCES 2010; 36(6): 474-482.
Bhui K, Tyagi S, Srivastava AK, Singh M, Roy P, Singh
Ahle NW, Hamlet MP. Enzymatic frostbite R, Shukla Y. Bromelain inhibits nuclear factor kappa-
eschar debridement by bromelain. Ann Emerg Med. B translocation, driving human epidermoid carcinoma
1987; 16(9): 1063-1065. A431 and melanoma A375 cells through G(2) /M arrest
Amini A, Moghaddam SM, Ehteda A, Morris L. to apoptosis. Mol Carcinogen. 2011; 51(3): 231-243.
Bromelain and N-acetylcysteine inhibit proliferation Bousser MG. Platelets, atherothrombosis, antiplatelet
and survival of gastrointestinal cancer cells in vitro: drugs and cerebral ischemia. B Acad Nat Med. 2013;
significance of combination therapy. J Exp 197(2): 389-394.
Clin Cancer Res. 2014; 33(1): 92. Buttner L, Achilles N, Bohm M, Hosseini KS, Mosges R.
Amini A, Masoumi-Moghaddam S, Morris DL. Efficacy and tolerability of bromelain in patients with
Bromelain. In: Utility of Bromelain and N- chronic rhinosinusitis--a pilot study. B-ENT. 2013;
Acetylcysteine in Treatment of Peritoneal 9(3): 217-225.
Dissemination of Gastrointestinal Mucin-Producing Cassano A, Drioli E, Galalaverna G, Marchelli R, Silvestro
Malignancies. Switzerland: Springer international R, Cagnasso P. Clarification and concentration of
publishing; 2016. p. 63-80. citrus and carrot juices by integrated membrane
Arshad ZIM, Amid A, Yusof F, Jaswir I, Ahmad K, Loke processes. J Food Eng. 2003; 57(1): 153-163.
SP. Bromelain: an overview of industrial application Chaanine AH, Hajjar RJ. AKT signalling in the failing
and purification strategies. Appl Microbiol Biot. 2014; heart. Eur J Heart Fail. 2011; 13(8): 825-829.
98(17): 7283-7297. Charles NS, Ward PA, Gilroy DW. Fundamentals of
Arumugam A, Ponnusami V. Pineapple fruit bromelain Inflammation. Yale J Biol Med. 2011; 84(1): 64-65.
recovery using recyclable functionalized ordered Chaurasiya RS, Sakhare PZ, Bhaskar N, Hebbar HU.
mesoporous silica synthesized from sugarcane leaf ash. Efficacy of reverse micellar extracted
Braz J Chem Eng. 2013; 30(3): 477-486. fruit bromelain in meat tenderization. J Food Sci
Asenjo JA, Andrews BA. Aqueous two-phase systems for Technol. 2015; 52(6): 3870-3880.
protein separation: a perspective. J Chromatogr A. Chen D, Huang S. Fast separation of bromelain by
2011; 1218(49): 8826-8835. polyacrylic acid-bound iron oxide magnetic
Babu BR, Rastogi NK, Raghavarao KSMS. Liquid–liquid nanoparticles. Process Biochem. 2004; 39(1): 2207-
extraction of bromelain and polyphenol oxidase using 2211.
aqueous two-phase system. Chem Eng Process. 2008; Chen T, Su S, Nie H, Zhu L. Display: Design and selection
47(1): 83-89. of functional binding peptides ligands application for
Baez R, Lopes MTP, Salas CE, Hernandez M. In isolation of bromelain using affinity chromatography.
Vivo Antitumoral Activity of Stem Pineapple J Biotechnol. 2008; 136(1): 620-632.
(Ananascomosus) Bromelain. Planta Med. 2007; Coelho DF, Silveira E, Pessoa JA, Tambourgi EB.
73(13): 1377-1383. Bromelain purification through unconventional
Bakhshaee M, Jabari F, Ghassemi MM, Hourzad aqueous two-phase system (PEG/ammonium
S, Deutscher R, Nahid N. The Prevalence of Allergic sulphate). Bioprocess Biosyst Eng. 2013; 36(2): 185-
Rhinitis in Patients with Chronic Rhinosinusitis. Iran J 192.
Otorhinolaryngol. 2014; 26(77): 245-249. Conrozier T, Mathieu P, Bonjean M, Marc JF, Renevier
JL, Balblanc JC. A complex of three natural anti-

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 Strategies to obtain bromelain for cure 13

inflammatory agents provides relief of osteoarthritis Graf J. Herbal Anti-Inflammatory Agents for Skin
pain. Altern Ther Health Med.2014; 20(1): 32-37. Disease. Skin Therapy Lett. 2000; 5(4): 3-5.
Corzo CA, Krzysztof, Waliszewski KN, Welti-Chanes J. Grzonka Z, Kasprzykowski F, and Wiczk W. Cysteine
Pineapple fruit bromelain affinity to different protein proteases. In: Polaina, J and P. MacCabe. Industrial
substrates. Food Chem. 2011; 133(3): 631-635. Enzymes, NY, USA: Springer; 2007.p. 181-195.
Darshan S, Doreswamy R. Patented anti- inflammatory Guo R, Canter PH, Ernst E. Herbal medicines for the
plant drug development from traditional medicine. treatment of rhinosinusitis: a systematic review,
Phytother Res. 2004; 18(5): 343-357. Otolaryn. Head Neck Surg. 2006; 135(4): 496-506.
De Lencastre NLC, Jozala AF, Lopes AM, De Carvalho Gupta R, Bradoo S, Saxena RK. Aqueous two-phase
SEV, Mazzola PG, Pessoa A. Stability, purification, systems: an attractive technology for downstream
and applications of bromelain: A review. Biotechnol processing of biomolecules. Curr Sci. 1999; 77(4):
Prog. 2016; 32(1): 5-13. 520-523.
Dhandayuthapani S, Perez HD, Paroulek A, Gupta YR, Gupta N, Rathi P. Bacterial lipases: an
Chinnakkannu P, Kandalam U, Jaffe M, Rathinavelu overview of production, purification and biochemical
A. Bromelain-induced apoptosis in GI-101A breast properties. Appl Microbiol Biotechnol. 2004; 64(6):
cancer cells. J Med Food. 2012; 15(4): 344-349. 763-781.
Dhaneshwar AD, Chaurasiya RS, Hebbar HU. Process Hale LP, Greer PK, Trinh CT, James CL. Proteinase
optimization for reverse micellar extraction of activity and stability of natural bromelain preparations.
stem bromelain with a focus on back extraction. Int Immunopharmacol. 2005; 5(4): 783-793.
Biotechnol Prog.2014; 30(4): 845-855. Hale LP, Greer PK, Trinh CT, Gottfried MR. Treatment
Doko B, Bassani V, Casadebaig J, Cavailles L, Jacob M. with oral bromelain decreases colonic inflammation in
Preparation of proteolytic enzyme extracts from the IL-10-deficient murine model of inflammatory
Ananascomosus L. Merr. fruit juice using semi bowel disease. Clin Immunol. 2005; 116(2): 135-142.
permeable membrane, ammonium sulfate extraction, Hale LP, Chichlowski M, Trinh CT, Greer PK. Dietary
centrifugation and freeze-drying processes. J supplementation with fresh pineapple juice decreases
Immunopharmaco. 2005; 4(5): 783-795. inflammation and colonic neoplasia in IL-10-deficient
Eckert K, Grabowska E, Stange R, Schneider U, mice with colitis. Inflamm Bowel Dis.2010; 16(12):
Eschmann K, Maurer HR. Effects of oral bromelain 2012-2021.
administration on the impaired immunocytotoxicity of Harrach T, Eckert K, Maurer HR, Machleidt I, Machleidt
mononuclear cells from mammary tumor patients. W, Nuck R. Isolation and characterization of two forms
Oncol Rep. 1999; 6(6): 1191-1199. of an acidic bromelain stem proteinase. J Protein
Errasti ME, Prospitti A, Viana CA, Gonzalez MM, Ramos Chem. 1998; 17(4): 351-361.
MV, Rotelli AE, Caffini NO. Effects on fibrinogen, Harrach T, Gebauer F, Eckert K, Kunze R, Maurer H.
fibrin, and blood coagulation of proteolytic extracts Bromelain proteinases modulate the cd44 expression
from fruits of Pseudananasmacrodontes, on human molt-4/8 leukemia and sk-mel-28
Bromeliabalansae, and B. hieronymi (Bromeliaceae) in melanoma-cells in-vitro. Int J Oncol.1994; 5(3): 485-
comparison with bromelain. Blood Coagul 488.
Fibrinolysis. Epub 2016, Feb 15. Hebbar HU, Hemavathi AB, Sumana B, Raghavarao
Ferreira FJ, Santana JCC, Tambourgi EB. The effect of KSMS. Reverse micellar extraction of bromelain from
pH on bromelain partition from Ananascomosus by pineapple (ananascomosus l. merryl) waste: scale-up,
PEG4000/Phosphate ATPS. Braz Arch Biol reverse micelles characterization and mass transfer
Technol. 2011; 54(1): 125-132. studies. Sep Sci Technol. 2011; 46(10): 1656-1664.
Fileti AM, Fischer GA, Tambourgi EB. Optimizing Hebbar HU, Sumana B, Hemavathi AB, Raghavarao
bromelain extraction by reversed micelles from KSMS. Separation and Purification of Bromelain by
pineapple fruit. Chem Eng Transc. 2007; 11(1): Reverse Micellar Extraction Coupled Ultrafiltration
989-994. and Comparative Studies with Other Methods. Food
Fileti AMF, Fischer GA, Santana JCC, Tambourgi EB. Bioprocess Technol.2012; 5(3): 1010-1018.
Batch and continuous extraction of bromelain Hebbar HU, Sumana B, Raghavarao KSMS. Use of
enzyme by reversed micelles. Braz Arch Biol reverse micellar systems for the extraction and
Technol. 2009; 52(5): 1225 -1234. purification of bromelain from pineapple wastes.
Gabrielli A, Avvedimento EV, Krieg T. Scleroderma. New Bioresource Technol. 2008; 99(1): 4896-4902.
Engl J Med. 2009; 360(19): 1989-2003. Helms S, Miller A. Natural treatment of chronic
Gaby AR. Natural remedies for scleroderma. Altern Med rhinosinusitis.Altern Med Rev. 2006; 11(3): 196-207.
Rev.2006; 11(3): 188-195. Hemavathi AB, Hebbar HU, Raghavarao KSMS. Reverse
Gautam SS, Mishra SK, Dash V, Goyal AK, Rath G. micellar extraction of bromelain from Ananascomosus
Comparative study of extraction, purification and L. Merryl. J Chem Tech Biotech. 2007; 82(11): 985-
estimation of bromelain from stem and fruit of 992.
pineapple plant. Thai J Pharm. 2010; 34(1): 67-76.

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 14 Nawaz, A et al.

Henriksson K, From J, Stratelis G. Patient-reported Krieger Y, Bogdanov-Berezovsky A, Gurfinkel

adherence to coprescribed proton pump inhibitor R, Silberstein E, Sagi A, Rosenberg L. Efficacy of
gastroprotection in osteoarthritis, rheumatoid arthritis, enzymatic debridement of deeply burned hands.
and ankylosing spondylitis patients using nonsteroidal Burns. 2012; 38(1): 108-112.
anti-inflammatory drugs. Patient Prefer Adherence. Kumar S, Hemavathi AB, Hebbar HU. Affinity based
2014; 8(1): 1611-1617. reverse micellar extraction and purification of
Hossain MM, Lee SI, Kim IH. Effects of bromelain bromelain from pineapple (Ananascomosus L. Merryl)
supplementation on growth performance, nutrient waste. Process Biochem. 2011; 46(5): 1216-1220.
digestibility, blood profiles, faecal microbial shedding, Larocca M, Rossano R, Santamaria M, Riccio P. Analysis
faecal score and faecal noxious gas emission in of pineapple [Ananascomosus (L.) Merr.] fruit
weanling pigs. Veterinarni Medicina. 2015; 60(10): proteinases by 2-D zymography and direct
544-552. identification of the major zymographic spots by mass
Hu W, Wang AM, Wu SY, Zhang B, Liu S, Gou YB, spectrometry. Food Chem. 2010; 123(1): 1334-1342.
Wang JM. Debriding effect of bromelain on firearm Lee KL, Chong CC. The use of Reverse Micelles in
wounds in pigs. J Trauma. 2011; 71(4): 966-972. Downstream Processing of Biotechnological
Hung TH, Chang YM, Sung HY, Chang CT. Products. 2011.
Purification and characterization of hydrolase with Ley CM, Tsiami A, Ni Q, Robinson N. A review of the use
chitinase and chitosanase activity from commercial of bromelain in cardiovascular diseases. J Chin Integr
stem bromelain. J Agric Food Chem. 2002; 50(16): Med. 2011; 9(7): 702-710.
4666-4673. Lightfoot EN. 1990. Protein purification from molecular
Jaber R. Respiratory and allergic diseases: From upper mechanisms to large scale processes. In: Ladisch MR,
respiratory tract infections to asthma. Prim Care. 2002; Willson RC, Painton CC, Builder SE. Separation in
29(2): 231-261. biotechnology-the key role of adsorptive separations.
Illanes A. Enzyme Production. In: Enzyme Biocatalysis. ACS Publications; 1990. p. 35-51.
Brazil: Springer Science and Business Media V.B. Lopes FLG, Junior JBS, Souza RR, Ehrhardt DD, Santana
2008; p. 57-106. JCC, Tambourgi EB. Concentration by Membrane
Johansson HO, Feitosa E, Pessoa A. Phase Diagrams of Separation Processes of a Medicinal Product Obtained
the Aqueous Two-Phase Systems of Poly (ethylene from Pineapple Pulp. Braz Arch Biol Technol. 2009; 52
glycol)/Sodium Polyacrylate/Salts. Polymers. 2011; (2): 457-464.
3(1): 587-601. MacKay ND, Miller AL. Nutritional Support for Wound
Juhasz B, Thirunavukkarasu M, Pant R, Zhan L, Healing. Altern Med Rev. 2003; 8(4): 359-377.
Penumathsa SV, Secor ER, Srivastava S,Raychaudhuri Manhart N, Akomeah R, Bergmeister H, Spittler A, Ploner
U, Menon VP, Otani H, Thrall RS, Maulik N. M, Roth E. Administration of proteolytic enzymes
Bromelain induces cardioprotection against ischemia- bromelain and trypsin diminish the number of CD4+
reperfusion injury through Akt/FOXO pathway in rat cells and interferon-gamma response in Peyer's patches
myocardium. Am J Physiol-Heart C. 2008; 294(3): and spleen in endotoxemicbalb/c mice. Cell Immunol.
1365-1370. 2002; 215(2): 113-119.
Jutamongkon R, Charoenrein S. Effect of Temperature on Marshall JS, Golden KD. Characterization of Bromelain
the Stability of Fruit Bromelain from Smooth Cayenne from Morindacitrifolia (Noni). J Sci Res. 2012; 4 (2):
Pineapple. Kasetsart J Nat Sci. 2010; 44(5): 943-948. 445-456.
Kaur H, Corscadden K, Lott C, Elbatarny HS, Othman M. Massimiliano R, Pietro R, Paolo S, Sara P, Michele F. Role
Bromelain has paradoxical effects on blood of bromelain in the treatment of patients with
coagulability: a study using pityriasislichenoideschronica. J Dermatolog Treat.
thromboelastography.Blood Coagul Fibrinolysis. 2007; 18(4): 219-222.
Epub 2014 Dec 16. Maulik, N, Das DK. Apoptosis, heart failure, ischemic
Ketnawa S, Chaiwut P, Rawdkuen S. Aqueous Two-phase heart disease. Heart Fail Rev. 1994; 4(1): 1-9.
Extraction of Bromelain from Pineapple Peels Maurer HR. Bromelain: biochemistry, pharmacology and
(‘PhuLae’ cultv.) and Its Biochemical Properties. Food medical use. Cell Mol Life Sci. 2001; 58(9): 1234-
Sci Biotechnol. 2011; 20(5): 1219-1226. 1245.
Ketnawa S, Rawdkuen S, Chaiwut P. Two phase Metzig C, Grabowska E, Eckert K, Rehse K, Maurer HR.
partitioning and collagen hydrolysis of bromelain from Bromelain proteases reduce human platelet
pineapple peel Nang Laecultivar. J Biochem Eng. aggregation in vitro, adhesion to bovine endothelial
2010; 52(2-3): 205-211. cells and thrombus formation in rat vessels in vivo. In
Ketnawa S, Sai-Ut S, Theppakorn T, Chaiwut P, Vivo. 1999; 13(1): 7-12.
Rawdkuen S. 2009. Partitioning of bromelain from Nadzirah KZ, Zainal S, Noriham A, Normah I. Efficacy of
pineapple peel (Nang Laecultv.) by aqueous two phase selected purification techniques for bromelain. IFRJ.
system. As J Food Ag-Ind. 2009; 2(4): 457-468. 2013; 20(1): 43-46.

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 Strategies to obtain bromelain for cure 15

Navapara RD, Avhad DN, Rathod VK. Application of Secor ER, Shah SJ, Guernsey LA, Schramm CM, Thrall
Response Surface Methodology for Optimization of RS. Bromelain limits airway inflammation in an
Bromelain Extraction in Aqueous Two-Phase System. ovalbumin-induced murine model of
Sep. Sci. Technol. 2011; 46(11): 1838-1847. established asthma. Altern Ther Health M. 2012; 18(5):
Neta JLV, DaSilva LA, Lima AA, Santana JC, Leite 9-17.
NS, Ruzene DS, SilvaDP,DeSouza RR. Bromelain Secor ER, Szczepanek SM, Castater CA, Adami AJ,
Enzyme from Pineapple: In Vitro Activity Study under Matson AP, Rafti ET, Guernsey L, Natarajan P,
Different Micropropagation Conditions. Appl Biochem McNamara JT, Schramm CM, Thrall RS, Silbart LK.
Biotechnol. 2012; 168(2): 234-246. Bromelain Inhibits Allergic Sensitization and Murine
Ng PK, He J, Synder MK. Separation of proteins mixtures Asthma via Modulation of Dendritic Cells. Evid Based
using PH-gradient cation exchange chromatography. J Complement Alternat Med. 2013; 2013(1): 1-9.
Chromatogr A. 2009; 1216(9): 1372-1376. Secor ER., Carson WF, Singh A, Pensa M, Guernsey LA,
Nie H, Li S, Zhou Y, Chen T, He Z, Su S, Zhang H, Xue Schramm CM, Thrall RS. Oral Bromelain Attenuates
Y, Zhu L. Purification of bromelain using immobilized Inflammation in an Ovalbumin-induced Murine Model
metal affinity membranes. J Biotechnol. 2008; 136: of Asthma. Evid Based Complement Alternat Med.
620-632. 2008; 5(1): 61-69.
Novaes LC, Ebinuma VC, Mazzola PG, Pessoa A. Shiraishi I, Melendez J, Ahn Y, Skavdahl M, Murphy E,
Polymer-based alternative method to extract bromelain Welch S, Schaefer E, Walsh K, Rosenzweig A, Torella
from pineapple peel waste. Biotechnol Appl Biochem. D, Nurzynska D, Kajstura J, Leri A, Sussman P.
2013; 60(5): 527-535. Nuclear targeting of Akt enhances kinase activity and
Onken JE, Greer PK, Calingaert B, Hale LP. Bromelain survival of cardiomyocytes. Circ Res.2004; 94(7): 884-
Treatment Decreases Secretion of Pro-Inflammatory 891.
Cytokines and Chemokines by Colon Biopsies. In Silvestre MPC, Carreira RL, Silva MR, Corgosinho FC,
Vitro ClinImmunol. 2008; 126(3): 345-352. Monteiro MRP, Morais HA. Effect of pH and
Orsini RA. Bromelain. Plast Reconstr Surg.2006; 118(7): temperature on the activity of enzymatic extracts from
1640-1644. pineapple peel. Food Bioprocess Technol. 2012; 5(5):
Pavan R, Jain S, Shraddha, Kumar A. Properties and 1824-1831.
Therapeutic Application of Bromelain: A Review. Singer AJ, McClain SA, Taira BR, Rooney J, Steinhauff
Biotechnol Res Int. 2012; 2012(1): 1-6. N, Rosenberg L. Rapid and selective
Pillai K, Akhter J, Chua TC, Morris DL. Anticancer enzymatic debridement of porcine comb burns
Property of Bromelain with Therapeutic Potential in with bromelain-derived Debrase: acute-phase
Malignant Peritoneal Mesothelioma. Cancer Invest. preservation of noninjured tissue and zone of stasis. J
2013; 31(4): 241-250. Burn Care Res.2010; 31(2): 304-309.
Rabelo APB, Tambourgi EB, Pessoa A. Bromelain Singer AJ, Taira BR, Anderson R, McClain SA,
partioning in twophase aqueous systems containing Rosenberg L. The effects of rapid
PEO-PPO-PEO block copolymers. J Chromatogr B. enzymatic debridement of deep partial-thickness burns
2004; 807(1): 61-68. with Debrase on wound reepithelialization in swine. J
Reddy KK, Grossman L, Rogers GS. Common Burn Care Res.2010; 31(5): 795-802.
complementary and alternative therapies with potential Smith-Marshall J. Golden KD. Characterization of
use in dermatologic surgery: Risks and benefits. J Am Bromelain from Morindacitrifolia (Noni). J Sci Res.
AcadDermatol. 2013; 68(4): 127-135. 2012; 4(2): 445-456.
Rosenberg L, Krieger Y, Silberstein E, Arnon Soares PAG, Vaz AFM, Correia MTS, Pessoa A Maria G,
O, Sinelnikov IA, Bogdanov-Berezovsky A, Singer Cunha C. Purification of bromelain from pineapple
AJ. Selectivity of a bromelain based wastes by ethanol precipitation. Sep Purif Technol.
enzymatic debridement agent: a porcine study. Burns. 2012; 98(2138): 389-395.
2012; 38(7): 1035-1040. Someshwar S, Udare S. Pityriasislichenoides. Indian
Rowan AD, Buttle DJ, Barrett AJ. The cysteine Pediatr.2012; 49(11): 936-941.
proteinases of the pineapple plant. Biochem J. 1990; Spir LG, Ataide JA, Novaes LC, Moriel P, Mazzola
266(3): 869-875. PG, Gurpilhares D, Silveira E, Pessoa A, Tambourgi
Saxena L, Iyer BK, Ananthanarayan L. Three phase EB. Application of an aqueous two-phase micellar
partitioning as a novel method for purification of ragi system to extract bromelain from pineapple
(Eleusinecoracana) bifunctional amylase/ protease (Ananascomosus) peel waste and analysis
inhibitor. Process Biochem.2007; 42(3): 491-495. of bromelain stability in cosmetic formulations.
Secor ER, Carson WF, Cloutier MM, Guernsey LA, Biotechnol Prog.2015; 31(4): 937-945.
Schramm CM, Wu CA, Thrall RS. Bromelain exerts Swaroop G, Viswanathan G. Isolation and
anti-inflammatory effects in an ovalbumin-induced Characterization of Bromelain (BML) Proteases from
murine model of allergic airway disease. Cell Immunol. Ananascomosus an asset to Cancer Chemotherapy.
2005; 237(1): 68-75. IJCPT. 2013; 1(2): 82-90.

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

 16 Nawaz, A et al.

Taussig SJ, Batkin S. Bromelain, the enzyme complex of Wu SY, Hu W, Zhang B, Liu S, Wang JM, Wang AM.
pineapple (Ananascomosus) and its clinical Bromelain ameliorates the wound microenvironment
application. An update. Ethno pharmacol. 1988; 22(2): and improves the healing of firearm wounds. J Surg
191-203. Res. 2012; 176(2): 503-509.
Taussig SJ, Szekerczes J, Batkin S. Inhibition of tumour Xue Y, Wu C, Branford-White CJ, Ning X, Nie H, Zhu L.
growth in vitro by bromelain, an extract of the Chemical modification of stem bromelain with
pineapple plant (Ananascomosus). Planta Med. 1985; anhydride groups to enhance its stability and catalytic
51(6): 538-539. activity. J MolCatal. B-Enzym. 2010; 63(3): 188-193.
Tochi BN, Wang Z, Xu SY, Zhang W. Therapeutic Yin L, Sun CK, Han X, Xu L, Xu Y, Qi Y, Peng J.
Application of Pineapple Protease (Bromelain): A Preparative purification of bromelain (EC 3.4.22.33)
Review. Pakistan J Nutrition. 2008; 7 (4): 513-520. from pineapple fruit by high-speed counter-current
Upadhyay A, Lama JP, Tawata S. Utilization of Pineapple chromatography using a reverse-micelle solvent
Waste: A Review. J Food Sci Technol. 2010; 6(1): 10- system. Food Chem. 2011; 129(3): 925-932.
18. Yuan G, Wahlqvist ML, He G, Yang M, Li D. Natural
Walker AF, Bundy R, Hicks SM, Middleton RW. products and anti-inflammatory activity. Asia Pac J
Bromelain reduces mild acute knee pain and improves ClinNutr. 2006; 15(2): 143-152.
well-being in a dose-dependent fashion in an open
study of otherwise healthy adults. Phytomedicine. Received: January 06, 2016;
Accepted: April 07, 2016.
2002; 9(8): 681-686.

Braz. Arch. Biol. Technol. v.59: e16150010, Jan/Dec 2016

View publication stats