Stem bromelain

Stem bromelain is a highly accepted phytotherapeutic agent. Muntari et al., 2012, reported that
bromelain concentration was very high in pineapple stems and hence leading to its extraction and
utilization as phytomedicinal compound. According to Ketnawa et al., 2012, there is a wide
range of applications of bromelain, hence a high demand of the enzyme. Thus, highly purified
commercial bromelain is not cheap; costing up to 2,400 USD per kilogram.
This review is aimed at presenting a detailed account on the level of achievements made on
stem bromelain production using the conventional methods. The extraction, purification and
biochemical properties of bromelain are considered.
Heinecke and Gotner (1957) reported that bromelain concentration was very high in pineapple
stems and hence leading to its extraction and utilization as phytomedicinal compound.
The commercial proteolytic enzyme preparations are evaluated according to their proteolytic
activity, which should be measured within the optimal conditions of enzymatic reactions.
Bromelain activity can be determined under optimal pH and temperature conditions, with various
substrates including casein, gelatin and synthetic substrates. From the several studies conducted,
Stem bromelain (SBM) was found have molecular weight range (26-37), pH optimum range (6-
7) and optimum temperature range of 50-60 º C (Harrach et al., 1998; Kumar et al., 2011; Suh et
al., 1992; Xue et al., 2010; Liang et al., 2011)

Properties of Stem Juice

The juice is a complex mixture of suspended material-starch granules, cell plastids, calcium
oxalate crystals, cellular debris and soil, and soluble material such as sugars, salts, complex
polysaccharides, proteins and lipids. Some of the properties of this juice are listed in Table 1.
(Heinecke and Gortner, 1957)

Table 1: Properties and composition of pineapple stem juice

* Milk clotting units are a measure of proteolytic activity (10).

† mM = Millimoles p-nitrophenol liberated from p-nitrophenylphosphate in one hour at pH 6.0 and 37.50 C.
‡: The absorbency at 400 mu of the color produced by the action of peroxidase and enzyme on a guaiacol solution at
pH 4.5 in 15 minutes at 37.50C.
Raw Material for the Enzyme and content
The highest concentration of proteases occurs in the lower portion of mature pineapple plant
stems. The central portion, the stele, contains more proteases than the outer portion, the cortex
(Figure 1). The less mature tissues, especially those that are still succulent, contain little or no
detectable proteases (Heinecke and Gortner, 1957).

Figure 2: Distribution of bromelain in various portions of the stem mature pineapple plant

Preparation of stem bromelain

Many of the classical precipitants can be used to prepare crude enzyme or certain fractions of
enzymes. Ammonium sulfate, methanol, isopropanol, and acetone combined with variations in
the pH of the juice have all been used in this investigation. However, for ease in preparing large
batches of bromelain, acetone is the most convenient precipitant. This is the precipitant which
has been employed on a pilot plant scale.
Acetone is added to the stem juices in two stages. When one to two volumes of acetone are
added to two volumes of stem juice, a precipitate forms which has low enzymatic activity, poor
color and very poor stability. This is discarded. Addition of another volume of acetone
precipitates the main enzymatic fraction. This is collected by centrifugation and dried. This dry
powder is stem bromelain. The acetone is recovered from the supernatant solution by distillation

Chemical Properties of Stem Bromelain

Stem bromelain prepared by acetone precipitation is a mixture of many colloids, inorganic salts
and simpler organic materials which are precipitated by acetone. Purre protein (trichloroacetic
acid insoluble N x 6.25) generally constitutes about 50% of the total weight
of dried precipitate. Inorganic materials, principally cations, generally make up 10-15% of the
total weight. The balance is assumed to be complex carbohydrate materials of the nature of
polyuronides and glycosides. Table 2 gives some of the materials in stem bromelain. The
complex carbohydrate materials are a major component of stem bromelain and affect the
physical properties of the mixture.

Table 2: Chemical composition of stem bromelain (Heinecke and Gortner, 1957)

Table 3: Various extraction and purification strategies for bromelain as employed by

different researchers
Table 4: The molecular weight, pH and temperature optima of bromelain as reported by
various research groups
Table 5: The different chemicals and strategies used for modification of bromelain in order
to enhance the stability of the enzyme
Table 6: Different immobilization techniques of bromelain for enhancement of its stability
and reusability
Table 7: Drying and stabilization of bromelain as conducted by various researchers
Figure 2: Schematic presentation of extraction and purification strategies of bromelain.
Several techniques that are being employed for the extraction and purification of
bromelain from different parts of pineapple plant (Bala et al., 2012)
Figure 3: Schematic presentation of biochemical characterization of bromelain. Different
techniques used by researchers for the characterization of bromelain in order to
standardized the enzyme (Bala et al., 2012)