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Plant secondary metabolism is an important source of fine chemicals such as drugs,

dyes, flavors, and fragrances. The technique to produce these chemicals is increasing

Pineapple plants is one of sources of proteases , most of the in vitro research on produce
protease from pineapple plants has been focused on propagation. like a procedure by
Escalona et al. (1999) involves the use of temporary immersion bioreactors.

The author previously found that pineapple shoots excrete proteases into the culture
medium during the pre-elongation phase of Escalona procedure

Therefore, The author decided to modify duration (15 d) and growth regulator
concentrations (2.8 mM GA and 2.2 mM BA) of the pre-elongation phase to increase
protease excretion into the culture medium

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1. Plant material : Pineapple buds

2. Culture medium: MS (Murashige and Skoog, 1962) salts, 100 mg l- 1 myo-inositol, 0.1
mg l - 1 thiamine-HCl, 30 g l - 1 sucrose, 4.4 µM 6-benzyladenine (BA), and 5.3 µM
naphthaleneacetic acid (NAA).

Cultures were maintained at 25 ± 1[C with 30 µE m- 2 s- 1 (fluorescent light) and an 8-

h photoperiod. Three bioreactors were sampled for evaluation every 7 d (0 28 d of culture).
The following indicators were recorded: shoot fresh mass per bioreactor; and protein
concentration, proteolytic activity, and specific activity in the culture medium
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· G. 1. a, Design of the temporary immersion bioreactor used in this paper (Escalona
et al., 1999). A, Air filter (0.2 mm) (Midsart 2000); B, glass vessel for shoots (300 ml); D,
silicone tube; E, glass vessel for liquid culture medium (250 ml); ·, liquid culture medium (250
ml); G, electric valve; H, air compressor. Shoots were free in the bottom of culture vessels (five
shoots per vessel). b, Mode of operation of a temporary immersion bioreactor. Steps 1, 2, and 3
were performed every 3 h. The air compressor and electric valves were under control of a
timer.
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