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110 วารสารการแพทย์แผนไทยและการแพทย์ ทางเลือก ปีที่ 20 ฉบับที่ 1 มกราคม-เมษายน 2565

mL of microorganisms. Microorganisms were evaluated using the MTT assay. Molt4 and
seeded onto Muller-Hinton agar (MHA) by us- KG-1a were seeded at a density of 1.0 5 10
4
ing a sterile cotton swab. Sterilized filter paper cells/well in 96-well plates and incubated

disks (Whatman no. 1; 6 mm) were individu- overnight at 37˚C with 5% of CO . Then, cells
2
ally impregnated with 10 μL of 200 mg/mL were treated with extract (0-100 μg/mL) for 48

extracts (resuspended in DMSO) to obtain 2 hours and the complete medium with DMSO
mg/disk. The filter paper discs impregnated was used as vehicle control (VC). After that
with the extracts were placed onto the seeded 15 μL of MTT dye (Sigma-Aldrich, St Louis,

plates. Ceftazidime 30 μg/disk was used as the MO, USA) solution (5 μg/mL) was added to
positive control. DMSO was used as negative each well and the plate was incubated at

control. The plates were incubated at 37˚C for 37˚C for 4 hours. Formed formazan crystals
24 hours. All tests were performed in triplicate were dissolved with 200 μL of DMSO, and the
and the antibacterial activity was expressed absorbance was measured at 578 nm by a Ac-

as mean of inhibition diameters (mm) pro- cuReader microplate reader (Metertech-Inc,
TM
duced. Taipei, Taiwan) and the reference blank was
2.8 cell line culturing and maintenance 630 nm. The % cell viability was calculated by

Molt4 and KG-1a were kindly provided by the formula as follows.
Dr. Songyot Anuchapreeda (Associate Profes- Absorbance of test 5 100
% Cell viability =
sor of Medical Technology within Faculty of Absorbance of control

Associated Medical Sciences at Chiang Mai 2.10 statistical analysis
University). Molt4 and KG-1a were used for All data were expressed as mean ± SE.

cytotoxicity assay. The cells were cultured in Analyses were performed in triplicates. The
modified RPMI-1640 complete medium with data were statistically evaluated using analysis
2.05 mM L-glutamine and 25 mM HEPES, of variance (ANOVA) with IBM SPSS Statis-

supplemented with 10% heat-inactivated FBS tics 28.0.0.0 version. Significance levels were
at 37˚C in a 95% humidified atmosphere with defined using p < 0.05.

5% CO . Cell counts and viability estimation by
2
trypan blue dye exclusion test were performed Results
regularly. Percentages of dead cells were in the

range of 0-3%. 1. Total phenolic contents
2.9 cytotoxicity assay The yield of L. leucocephala ethanolic
The cytotoxicity of crude extract was extract was about 25.85%. Estimation using