Evaluation of Cytotoxicity of Different Concentration of van Tulsi (Ocimum Gratissimum) on Roots Tip Cells Onion (Allium Cepa)

: The present study aimed to assess the cytotoxic effects of Van Tulsi (Ocimum gratissimum) leaf extract at a 5% concentration on root tip cells of Onion (Allium cepa) after exposure periods of 72 hours and 96 hours. Fresh Onion (Allium cepa) root tips were exposed to Van Tulsi leaf extract at a concentration of 5% for two different time intervals, 72 hours and 96 hours. Control groups with untreated root tips were also maintained for comparison. The cytotoxicity was evaluated through various cytological parameters, including cell viability, mitotic index-Prophase, Metaphase, Anaphase, Telophase. The findings revealed a significant decrease in cell viability in the treated groups compared to the control at both 72 hours and 96 hours of exposure. Additionally, the mitotic index was markedly inhibited in root tip cells exposed to Van Tulsi extract. The treated cells exhibited an elevated frequency of chromosomal aberrations and micronuclei, indicating genotoxic effects of the extract. The results of this study demonstrate that Van Tulsi (Ocimum gratissimum) leaf extract at a concentration of 5% exerts cytotoxic effects on root tip cells of Onion (Allium cepa). These findings contribute to our understanding of the effects of Van Tulsi leaf extract on plant cells and warrant further investigation into its underlying mechanisms of cytotoxic action. In 72 hrs treatment duration mitotic index slightly increased but at 96 hrs duration mitotic index significantly decreased due to decrease in the proportion of prophase and Metaphase cell population. Thus, leaf extract of Van Tulsi act as mitotic inhibitor on onion root tip cells at higher treatment duration.

The present study was carried out to evaluate the cytotoxic effect of Van Tulsi (Ocimum gratissimum) leaf extract on root tip cells of Onion (Allium cepa).

MATERIALS AND METHODS:
PREPARATION OF LEAF EXTRACT: The leaves of Van Tulsi (Ocimum gratissimum) were washed under running tap water and shade dried for 2 to 3 weeks. After that, dried fresh Tulsi leaves were homogenised by using a grinder to made fine powder so obtained and stored in air tight bottles. 100gram fine powder were dissolved in 1000ml distilled water as a stock solution and left for 48hours. It was then filtered through Whatman No. 1 [21]. Van Tulsi leaf extract 5% dose was prepared by dilution of stock solution [22].
The onion bulb weighing approximately 20-30 grams were purchased from local market and their roots were initially allowed to grow till 1.5 cm in length in normal tap water. The bulb roots were cut after 72hrs and 96hrs and fixed in aceto-alcohol for 24hrs then preserved in 70% ethanol and used as control group.
Another set of onion bulbs (20-30gm) were grown in 5% Van Tulsi leaf extract for 72hrs and 96hrs respectively and used as treated group.

SLIDE PREPARATION:
After treatment, slides were prepared by Acetocarmine squash preparation [23]. Approximately 4000 cells were randomly analyzed in both control and treated group of onion bulbs. Frequency of Mitotic index and Phases distribution were calculated.

SLIDE SCREENING:
All the slides were examined under light microscope. The mitotic index was calculated for determination of cytotoxicity. Mitotic index (MI) was calculated as the ratio between the number of mitotic cells and the total number of cells scored and expressed as percentage and represented by following formulae [23].

STATISTICAL ANALYSIS:
The data are expressed as Mean ± SE and statistical analysis was performed by using t-test.

RESULT AND DISCUSSION:
In 72hrs treatment duration there is no significant difference were found in mitotic index and phase distribution of Van Tulsi treated onion root tips cells than control group of onion root tips cells. This suggest that 72hrs concentration of Van Tulsi could not induce cytotoxicity.
In 96hrs treatment duration, the mitotic index significantly decreased from 34.63% to 22.00% and in phase distribution mitotic index of Prophase and Metaphase significantly decreased from 29.42 to 18.7% and 3.26 to 2.10%. This decrease in mitotic index was mainly due to a decrease in the population of cells belonging to Prophase and Metaphase (Table 1).
Similar results were observed [23]. Numerous studies have shown that wherever there is root growth inhibition in Allium test, there is also reduction in the number of dividing cells [24].