AN IN VIVO STUDY OF THE INFLUENCE OF ETHANOLIC LEAF EXTRACT

OF KATSUBONG (DATURA METEL VAR. FASTUOSA L.) ON PLATELET

COUNT AND OTHER HEMATOLOGICAL PARAMETERS OF
ALBINO MICE (MUS MUSCULUS)

PSYCHOLOGY AND EDUCATION: A MULTIDISCIPLINARY JOURNAL

Volume: 41
Issue 7
Pages: 796-801
Document ID: 2025PEMJ3998
DOI: 10.70838/pemj.410702
Manuscript Accepted: 05-11-2025
 Psych Educ, 2025, 41(7): 796-801, Document ID:2025PEMJ3998, doi:10.70838/pemj.410702, ISSN 2822-4353
Research Article

An In Vivo Study of the Influence of Ethanolic Leaf Extract of Katsubong (Datura Metel Var. Fastuosa
L.) On Platelet Count and other Hematological Parameters of Albino Mice (Mus Musculus)
Kristel Cassandra Remedios,* Sara Nhelen T. Biag, Patrizia Angela P. Claudio,
Mariegold Ann J. Japitana, Alfie A. Gardose
For affiliations and correspondence, see the last page.
Abstract
Datura metel var. fastuosa L. is traditionally used for its anti-inflammatory and analgesic properties, yet its effects on
hematological parameters and toxicity remain underexplored. This quantitative study evaluated the impact of
Katsubong ethanolic leaf extract on platelet count, red blood cell (RBC) count, white blood cell (WBC) count, and
hemoglobin levels in mice. A Completely Randomized Design (CRD) was employed using male albino mice treated
with extract doses of 0.1 mL, 0.2 mL, and 0.3 mL. Hematological parameters were measured pre- and post-treatment
using a veterinary hematology analyzer. The extract appeared to increase platelet counts in a dose-dependent manner,
with the 0.1 mL dose showing slight improvement, 0.2 mL producing a more noticeable increase, and 0.3 mL showing
the most significant rise, particularly in R2 and R3. Despite this trend, the control and treatment groups for pre- and
post-extraction platelet counts revealed a mean increase from 161.78 to 385.89. However, the two-tailed p-value of
0.143 and the t Stat of -2.349 in the paired two-sample t-test were above the 0.05 significance level. Thus, the null
hypothesis was accepted, indicating no statistically significant difference in platelet counts. Hemoglobin levels
improved across all treatments, with Treatments 1 and 2 showing the greatest increases. The WBC counts rose in
Treatments 1 and 3, while RBC counts varied, with a decrease observed in Treatment 2. For toxicity testing, the Brine
Shrimp Lethality Assay (BSLA) was conducted by exposing Artemia salina nauplii to varying extract concentrations,
and LC₅₀ values were interpreted using Clarkson’s criteria. Results showed that the Katsubong extract exhibited low
toxicity. These findings suggest Katsubong extract may influence platelet and hemoglobin levels. However, the
observed inconsistencies and lack of statistical significance highlight the need for further studies on its therapeutic
potential.
Keywords: concentrations, datura metel var. fastuosa l., platelet, white blood cells, red blood cells, in vivo, albino
mice, extraction, hematological parameters

Introduction
Datura plant parts, extracts, and preparations have been used as medicines by humans for millennia. Immune thrombocytopenia and
other conditions, such as certain viral infections, bone marrow disorders, or autoimmune diseases, can lead to decreased platelet counts.
(McCrae, 2017). However, while most studies concentrate on Datura's toxicity and pharmacological activities, its potential on
hematology, specifically platelet function, remains widely unexplored. The history on the investigation of platelet function began 100
years ago, and thrombocytopenia is one of the most frequent causes for hematologic consultation in the practice of medicine and may
be life threatening (Kim, 2022). Consequently, investigating the potential of Datura metel var. Fastuosa L. in platelet count may
contribute to possible treatment or alternative medicine.
For decades, humans have been using extracts from plant materials for the treatments of many diseases. Plants have always been a
good source of biologically active compounds. Therapeutic plants are an essential element of ethnomedicine for both human and animal
uses all over the world. They have been in use for centuries to treat disease and improve health. On the other hand, one major concern
for modern medical practitioners regarding medicinal plants is their safety. Researchers have employed various laboratory assays to
assess the safety of herbal plants and products derived from them, but challenges still persist (Clemen-Pascual, 2021). Toxicity tests
are essential for assessing the lethality of substances and determining the safe concentration of drugs for medicinal use. One commonly
employed method for testing the toxicity of plant materials is the brine shrimp (Artemia sp.) lethality assay (BSLA). The advantage of
using brine shrimp in toxicity testing lies in the high homogeneity of their eggs and newly hatched nauplii, which are highly sensitive
to chemicals. Additionally, this method requires only a small amount of test material (Sharna, 2021). Furthermore, the brine shrimp
lethality assay is a valid indicator of the harmful potential of natural product extracts in humans, as evidenced by the positive correlation
between Meyer's toxicity scale for Artemia salina and Gosselin, Smith, and Hodge's toxicity scale for higher animal models (Osman
& Omar, 2019).
Mice play an essential role in medical research due to their close biological and genetic similarities to humans. Researchers recognized
early on that studying mice can offer valuable findings into various medical conditions, such as cancer and rare diseases (Ewumi,
2023). Despite their outward differences from humans, mice share around 95% of their genes with humans, making them highly
effective models for human biology. Many diseases in mice are genetically similar to those in humans, allowing for deeper
understanding of shared disease mechanisms (Max-Planck-Gesellschaft, 2024). Furthermore, in vivo testing is an essential part of
medical research in general, particularly in clinical trials. Reliable information on the effects of a specific chemical or the course of a
disease in a full, live organism may be obtained through in vivo research (Eske, 2023).
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Research Article

This study aims to investigate the potential of Datura metel var. Fastuosa L. as an alternative medicine for increasing platelet countA.
Furthermore, this study will assess its effectiveness on other hematological parameters such as white blood cells, red blood cells, and
hemoglobin levels. The researchers also recognized the risks associated with Datura metel var. fastuosa L. given the toxic profile of its
neighbouring variety and therefore will conduct a toxicity assessment for a safe approach. Awareness of diseases like thrombocytopenia
frequently require medical consultation, the researchers aim to determine if compounds found in Datura metel may promote or improve
platelet function. To address the research gap, this study will utilize in vivo testing with albino mice to thoroughly investigate the
toxicity and hematological effects of Datura metel. Therefore, the study seeks to provide significant findings that could pave the way
for developing new, effective treatments for low platelet count, or thrombocytopenia.
Methodology
Research Design
A Completely Randomized Design (CRD) was utilized in this study. It consists of 3 treatments replicated 3 times. It is widely used in
laboratory-based biological and toxicological research. CRD is most effective when experimental units such as lab animals or biological
models which are considered reasonably homogeneous. It minimizes bias by ensuring each unit has an equal chance of receiving any
treatment, thus enhancing the reliability of results (Sinba, 2010). In medical and toxicological studies, its randomized nature enables a
more objective evaluation of the effects of various treatments and interventions (Costello, 2023). Its simplicity and effectiveness in
ensuring equal chance of receiving any treatment make it appropriate for this study, which involved both in vivo application on mice
and in vitro toxicity testing using the Brine Shrimp Lethality Assay (BSLA).
In Vivo application and Brine Shrimp Lethality Assay:
T1- 0.1ml of katsubong ethanolic leaf extract
T2- 0.2ml of katsubong ethanolic leaf extract
T3- 0.3ml of katsubong ethanolic leaf extract.
Procedure
Prior to experimentation, ethical standards were followed such as ethical clearance obtained from the local City Veterinary Office and
oversight provided by a licensed veterinarian. All test subjects were housed in well-ventilated cages and monitored regularly. Extract
dosages were administered based on the veterinarian’s recommendations to ensure they were within tolerable limits and caused no
unnecessary harm. All procedures followed the principles set forth in the International Guiding Principles for Biomedical Research
Involving Animals (CIOMS, 2012), emphasizing animal welfare, scientific justification, and responsible care.
Data were gathered through direct measurements. Toxicity was assessed using the Brine Shrimp Lethality Assay (BSLA), where twenty
(20) brine shrimp nauplii were exposed to varying concentrations (0.1 mL, 0.2 mL, and 0.3 mL) of Datura metel extract. Mortality was
monitored and recorded after 24 hours using a microscope for accuracy. For the hematological analysis, platelet counts were measured
using a veterinary hematology analyzer. Blood samples were collected from mice before and 24 hours after subcutaneous administration
of the plant extract at the same concentrations. These data were used to determine the extract's effect on platelet levels.
Data from the Brine Shrimp Lethality Assay (BSLA) were processed using Microsoft Excel, with Probit analysis used to determine the
LC₅₀ or median lethal concentration. The researchers used the toxicity indices developed by Meyer et al. (1982) and Clarkson et al.
(2004) to interpret these results. To assess the extract’s effect on platelet count, a paired two-sample t-test was performed comparing
pre- and post-treatment values.
Data Analysis
The data gathered from the Brine Shrimp Lethality Assay (BSLA) were processed using Microsoft Excel, and Probit analysis was
applied to calculate the LC₅₀ values, which indicate the median lethal concentration of the extract. Additionally, a paired two-sample
t-test was conducted to compare the hematological parameters between the pre-extraction and post-extraction phases, allowing for the
assessment of significant differences attributable to the treatment. This combination of statistical tools provided a comprehensive
evaluation of both toxicity levels and physiological effects.
Ethical Considerations
This study ensured full ethical compliance throughout all stages of the research. The welfare of the animal subjects was prioritized by
adhering to humane handling protocols, with ethical clearance obtained from the local City Veterinary Office and oversight provided
by a licensed veterinarian. All test subjects were housed in well-ventilated cages and monitored regularly. Extract dosages were
administered based on the veterinarian’s recommendations to ensure they were within tolerable limits and caused no unnecessary harm.
All procedures followed the principles set forth in the International Guiding Principles for Biomedical Research Involving Animals
(CIOMS, 2012), emphasizing animal welfare, scientific justification, and responsible care. Additionally, data confidentiality and
integrity were strictly maintained throughout the research process.

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Research Article

Results and Discussion

Figure below shows the toxicity of Katsubong (Datura metel var. fastuosa L.). Using the Brine Shrimp lethality Assay (BSLA)

Figure 1. Toxicity of Katsubong (Datura metel var. fastuosa L.)

The researchers used the toxicity indices developed by Meyer et al. (1982) and Clarkson et al. (2004) to interpret these results. Herbal
extracts having an LC50 value less than 1000 μg/mL are categorized as toxic, whereas those with values greater than 1000 μg/mL are
regarded as non-toxic, using Meyer's toxicity index. Katsubong extract was classified as hazardous under these guidelines. However,
Clarkson's toxicity categorization adds further subtlety by classifying LC50 values as mild toxicity if they fall between 500 and 1000
μg/mL. With an LC50 value of 626.62 μg/mL, Katsubong extract falls within Clarkson's low toxicity category, indicating minimally
hazardous effects at moderate dosages. Higher extract concentrations resulted in higher mortality rates for the brine shrimp Nauplius
mortality percentages were 0% for 1 μg/mL, 1.67% for 100 μg/mL, 6.67% for 500 μg/mL, 11.67% for 1000 μg/mL, and 35% for 2000
μg/mL. The toxicity level of Katsubong extract is quantitatively measured by its LC50 value, which was determined to be 626.62
μg/mL. These results indicate that although Katsubong extract is poisonous at greater dosages, it may be harmless in some measured
conditions. According to other studies, Datura metel contains bioactive chemicals that may be medicinal but should be handled carefully
to prevent adverse effects. Additional research on the precise substances causing toxicity would help produce safe dosage guidelines
and offer significant insights into Katsubong's safe therapeutic range. This shows how important it is to achieve balance between safety
and effectiveness, especially when using herbal remedies made from species that are known to have strong bioactive ingredients
(Katoch, et. al, 2024).
Table 1. Effects and Notable Mean Differences in Platelet Count Following Administration of ethanolic Leaf Extract of Katsubong
(Datura metel var. fastuosa L.)
Treatments Replicates
R1 R2 R3
before after mean before after mean before after mean difference
difference difference
T1- 0.1ml Katsubong (Datura 571 609 38 33 191 158 37 180 143
metel var. fastuosa L.) leaves
extract
T2- 0.2ml Katsubong (Datura 37 122 85 106 174 68 104 683 579
metel var. fastuosa L.) leaves
extract
T3- 0.3ml Katsubong (Datura 179 197 18 283 1010 727 106 603 497
metel var. fastuosa L.) leaves
extract

The ethanolic leaf extract of Katsubong (Datura metel var. fastuosa L.) appears to influence platelet counts differently depending on
the concentration used. At the lowest dose (0.1 mL), there was a slight increase in platelet counts across all trials, suggesting a mild
effect on platelet production. The 0.2 mL dose showed a more noticeable improvement, particularly in one trial (R3), indicating that
this concentration might be more effective in boosting platelet levels. At the highest dose (0.3 mL), platelet counts increased even more
significantly, especially in R2 and R3, suggesting that the extract's effects become stronger as the dose increases. This work highlights
how specific treatments or factors can induce a progressive increase in platelet count, often correlating with the administered dose or
intervention's potency (Shivdasani, 2001).

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Research Article

Table 2. Notable Mean Differences between Control and Treatments in Pre- and Post-
Extraction Platelet Counts Using Ethanolic Leaf Extract of Katsubong (Datura metel
var. fastuosa L.)
Variable 1 Variable 2
Mean 161.7767 385.89
Variance 4881.939 37910.37
Observations 3 3
Pearson Correlation 0.569238
Hypothesized Mean Difference 0
df 2
t Stat -2.34917
P(T<=t) one-tail 0.071633
t Critical one-tail 1.885618
P(T<=t) two-tail 0.143266
t Critical two-tail 2.919986

Comparing the control and treatment groups for pre- and post-extraction platelet counts of the ethanolic leaf extract of Katsubong
(Datura metel var. fastuosa L.) revealed a mean increase from 161.78 to 385.89. However, the two-tailed p-value of 0.143 and the t
Stat of -2.349 in the paired two-sample t-test were higher above the significance level of 0.05. Therefore, the null hypothesis is accepted,
showing that there is no statistically significant difference in platelet counts as measured. To confirm the extract's ability to raise platelet
count, more research with a bigger sample size and higher statistical power is required. Emphasizing that statistical insignificance may
reflect limited sample size or variability and does not necessarily rule out meaningful findings. Additional replication or larger samples
are often needed to validate such results. (Andrade, 2019).
Table 3. Significant Difference between the Control and Treatments in Pre- and Post-Treatment Platelet Counts
Following the Administration of Ethanolic Leaf Extract of Katsubong.
Treatments Treatments Control
(before) (after) mean difference (before) (after) mean difference
T1 213.67 326.67 113 550 335 215
T2 82.33 227.67 145.34 208 89 199
T3 189.33 603.33 414 336 379 43

Effects of the ethanolic leaf extract of Katsubong on platelet counts by comparing pre- and post-treatment values across three treatment
groups and a control. In T1, platelet counts increased moderately, with a mean difference of 113. T2 showed a greater improvement,
with a mean difference of 145.34, while T3 exhibited the most significant increase, with platelet counts rising by 414. In comparison,
the control groups experienced minimal changes, with platelet counts decreasing slightly in T2 by 199 and increasing marginally in T3
by 43. These findings indicate a dose-dependent relationship, where higher concentrations of the extract lead to greater increases in
platelet counts, highlighting its potential therapeutic benefits. Katsubong can enhance platelet counts more effectively than prednisone
due to their bioactive compounds. In contrast, prednisone works by suppressing immune responses, which may result in slower or less
pronounced increases in platelet counts. Additionally, individual variations in responses to prednisone may limit its effectiveness.
(Aird, 2024).
Table 4. Effects of ethanolic Leaf Extract of Katsubong (Datura metel var. fastuosa L.) on Hematological Parameters: Hemoglobin,
Red Blood Cells, and White Blood Cells
Treatments Hemoglobin Red blood cells White blood cells
before after mean difference before after mean difference before after Mean difference
T1 2.67 3.72 1.05 1.67 2.33 0.66 1.67 4.14 2.47
T2 2.33 3.38 1.05 6.33 1.33 5.0 5.37 5.33 0.04
T3 1.50 2.70 1.02 0.66 1.67 1.01 1.67 3.98 2.31

The ethanolic leaf extract of Katsubong (Datura metel var. fastuosa L.) demonstrated varying effects on hematological parameters,
including hemoglobin, red blood cells (RBC), and white blood cells (WBC). Hemoglobin levels increased across all treatments, with
T1 and T2 showing the highest mean increases of 1.05, while T3 followed closely with a mean increase of 1.02. For RBC, T1 and T3
exhibited moderate increases with mean differences of 0.66 and 1.01, respectively, but T2 showed a significant decrease, with a mean
reduction of 5.0. In terms of WBC, T1 and T3 displayed notable increases, with mean differences of 2.47 and 2.31, while T2 showed
only a minimal change of 0.04. These results suggest that the Katsubong extract may positively influence hemoglobin and WBC levels,
particularly in T1 and T3, though its effects on RBC are inconsistent across treatments. Interventions can lead to improvements in some
hematological parameters, such as hemoglobin or white blood cells, while potentially causing reductions in others, depending on the
treatment conditions and individual responses. individual responses (Delaney et al., 2011).
Conclusions
This study highlights the potential of the ethanolic leaf extract of Datura metel var. fastuosa L. (Katsubong) as a plant-based agent with

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Research Article

hematological effects. Although the extract exhibited low toxicity based on LC₅₀ results, increasing concentrations led to higher brine
shrimp mortality, emphasizing the importance of safe dosage regulation. The extract showed trends of improving platelet count and
hemoglobin levels, though the changes were not statistically significant. These preliminary findings point to the possibility of
therapeutic value, particularly in enhancing certain blood parameters.
However, the inconsistent effects on red blood cell counts and the absence of significant statistical outcomes suggest that conclusions
on its efficacy remain inconclusive. These results underscore the need for more comprehensive research, including compound isolation,
mechanistic studies, and larger-scale trials, to better understand its pharmacological potential and ensure its safety for possible
therapeutic use.
In light of the study’s outcomes, it is recommended that future investigations: (1) conduct in vitro assays to elucidate the extract’s
specific biological actions; (2) perform phytochemical analyses to identify the compounds contributing to its toxicity and hematological
effects; (3) assess the impact of oral administration to simulate realistic therapeutic use; (4) expand the range and depth of hematological
parameters examined; and (5) increase the number of replicates to enhance statistical power and generalizability. These directions are
essential to inform safe and evidence-based applications of Datura metel var. fastuosa L. in pharmacological or clinical settings.
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Affiliations and Corresponding Information
Kristel Cassandra Remedios
Tacurong National High School
Department of Education – Philippines
Sara Nhelen T. Biag
Tacurong National High School
Department of Education – Philippines
Patrizia Angela P. Claudio
Tacurong National High School
Department of Education – Philippines

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Research Article

Mariegold Ann J. Japitana

Tacurong National High School
Department of Education – Philippines
Alfie A. Gardose
Tacurong National High School
Department of Education – Philippines

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