Page 145 - phytochemistry general program

precisely determined during manufacturing. A filter of pore size 0.22 µm is enough
to eliminate the smallest bacteria.

d- Chemical sterilization

The working area is generally surface-sterilized with either ethanol or isopropanol
(70% v/v). This method is also used for the sterilization of the plant material. It
should be surface sterilized to remove microorganisms with a minimum damage to
the plant system to be cultured. High concentration or longer contact time with the

reagent may reduce percentage of germination.

Surface sterilization will be ineffective if plant tissue contains microorganisms
within the tissue. If fleshy organs are selected for explant preparation, any
localized internal discoloration should be removed. An aqueous solution of sodium
hypochlorite (NaOCl) or calcium hypochlorite (Ca(OCl)2) are used for the surface
sterilization of a plant material. House hold bleaching agent such as clorox or

purex can be used.

The final sterilization solution should contain 0.5% of the active agent (Ca(OCl)2)
solution is preferred than NaOCl, because sodium ions may cause abnormal
development (due to its small ionic size). The explant is immersed in the
hypochlorite solution for 10 minutes, if the plant contain cutin, suberin or hairs a
wetting agent such as tween 80 and teepol are used, after that the plant should be
washed several times with sterile distilled water to remove any traces of the

hypochlorite solution.

e-Antibiotics

Although some workers have supplemented nutrient media with antibiotics for the
control of microbial contamination, these complex natural products are not

recommended now because of the following:

1- They may alter the growth and development of P.T.C. in vitro.

2- There is no antibiotic which is effective against all possible microorganisms
which may cause contamination.

3- These products may be metabolized by the plant tissue with unexpected results.

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