Materials and Methods

Preparation of Crude Extract from Butterfly Pea Flowers

Butterfly pea petals were air-dried in the shade and ground into a fine powder. The
powder was gradually dissolved in distilled water at a ratio 1:10 at room
temperature. The crude extract was collected by initially filtering with gauzes and
subsequently with No. 1 Whatman grade filter papers (Whatman, 1001-150). The pH
of the crude extract was measured using a CyberScan 1000 pH meter (Eutech
Instruments Pte Ltd, Singapore).
Preparation of Butterfly Pea Dye for Staining
The filtered crude extract of butterfly pea petals was added to 1.0% aluminium
chloride anhydrous and 1.2% Iron (III) chloride hexahydrate. The dye solution
wasmixed well and filtered using No. 1 Whatman grade filter papers. The pH was
adjusted to 0.2 using concentrated HCl.
Staining on Blood Smear
The staining process was designed to be conducted on blood smears of chicken,
pigeon, dog, and horse. The smears were prepared from EDTA-blood on glass slides.
The slides were air-dried and fixed in Diff-quick fixative reagent which contained
methanol and triarylmethane dye for 30-60 sec. The slides were stained with the
butterfly pea dye for 30 min. The staining process was stopped by covering the
slides with glass covers without washing and counter-staining. Images of the stained
smears were taken under an Axiolab microscope using Axio Vision software. All of
the process was conducted under light-protected conditions.
http://www.thaiscience.info/journals/Article/SJST/10890421.pdf
 TITLE Utilization of an indigenous dyestuff from Basella rubra
(alugbati) as microbiological stain

CALL NO(S) Fil Q149.P5 N25

LOCATION(S) STII

Transactions of the National Academy of Science and

PUBLICATION TITLE
Technology

VOLUME/ISSUE 28(1)

ISSUE DATE 2006

MAIN AUTHOR Enerva, Lorna T. , et al

This study was undertaken to extract anthocyanin from

Basella rubra berries and utilize the extract as microbiological
stain. It is an inexpensive, indigeneous and abundant raw
material. The alugbati berries were macerated in a blender and
extracted with 1% HCl in 95% methanol. The extract obtained
was filtered and then concentrated. Thin layer and column
chromatography methods were used to isolate and purify the
anthocyamin. The samples were analyzed using infrared spectra
and ultraviolet spectra. FT-IR revealed the presence of a
hydroxyl group which is prominent in the structure of
anthocyanin pigment at 3385cm'1. The C=O stretching for
aromatic ring were indicated by a peak at 1635.20 cm'l,
1513.38 em-I for C=O bending, 1439.91 cm'I for O-H bending,
1207.42 cm,l for c-o stretching, 1151.92cm" for alkane and
1100.77 em-I for C-C stretching. The structure of anthocycanin
ABSTRACT
was further established by the e maximum of the ultraviolet
spectrum at 510.0 nm. For the application, the crude extract
was used as a stain for Staphyloccus aureus, a gram positive
bacteria and Escherichia coli, a gram negative bacteria. The
staining process for the microorganism used mordants like
potassium alum, calcium oxide and copper sulfate for fixing the
color. Only copper sulfate and lime responded positively as a
mordant that gave favorable outcome in fixing the color of
alugbati. The samples were screened based on the criteria of
color retention and evenness. The structure of the
microorganisms with respect to shape and size and certain
cellular components were identified using a microscope and
photomicrographs. The alugbati extract produced stain that was
comparable with synthetic stains like crystal violet and safranin
and can, therefore, be used as an alternative stain.

SUBJECTS Biological sciences

Basella rubra (alugbati)
Microbiological stain