UNIVERSA MEDICINA
pISSN: 1907-3062 / eISSN: 2407-2230 Univ Med 2025.
44:172-180
DOI: https://doi.
org/10.
18051/UnivMed.
ORIGINAL ARTICLE
Hepatoprotective effects of celery in diethylene glycol induced toxicity in rats Heny Ekowati .
Syaikah Syahidah Waluyo , and Nialiana Endah Endriastuti Department of Pharmacy.
Faculty of Health Sciences.
Jenderal Soedirman University.
Purwokerto.
Indonesia * Correspondence Author:
ekowati@unsoed.
Date of first submission.
March 2, 2025 Date of acceptance.
July 25, 2025 Date of publised.
August 4, 2025 Cite this article as: Ekowati H.
Waluyo SS.
Endriastuti NE.
Hepatoprotective effects of celery in diethylene glycol induced toxicity in rats.
Univ Med 2025.
44:172-180
ABSTRACT
BACKGROUND
Diethylene glycol (DEG) is a contaminant in pharmaceutical solvents and potentially induces hepatotoxicity.
Celery is hepatoprotective due to its antioxidant properties.
The objective of this study was to evaluate the hepatoprotective effects of celery on DEG-induced liver injury in rats.
METHODS
An experimental laboratory study was conducted involving 25 male Wistar rats weighing 200 grams.
They were randomized into five groups .
rats in each grou.
Group A served as the control group, while Group B received oral administration of DEG at a dose of 3 g/kg BW twice daily for three days, following six days of food and water administration.
The other three groups received DEG at 3 g/kg BW following six days of celery extract once daily administration of 100 mg/kg BW (C), 200 mg/kg BW (D), and 400 mg/kg BW (E).
After 14 days, all rats were sacrificed.
Observations were conducted macroscopically and microscopically using H&E staining.
Liver damage was scored.
Kruskal-Wallis followed by the Mann-Whitney was used to analyze the data.
RESULTS
Macroscopic and microscopic examinations of the liver tissue in the celery groups revealed normal histological architecture with a significant reduction in liver damage.
The ethanol extract of celery demonstrated hepatoprotective effects, as evidenced by a statistically significant reduction .
<0.
in the activation of Kupffer cells, sinusoidal dilation, hepatocyte degeneration, and necrosis.
CONCLUSION
Celery has a protective effect against DEG-induced liver damage in rats based on the reduction in Kupffer cell activation, sinusoidal dilation, hepatocyte degeneration, and necrosis.
Keywords: Celery, hepato-protector, diethylene glycol, rats Copyright@Author.
- https://univmed.
org/ejurnal/index.
php/medicina/article/view/1724 Effect of celery in DEG toxicity INTRODUCTION The liver serves as the principal organ for detoxification within the human body, playing a critical role in the metabolism of pharmaceuticals and toxic substances.
The majority of drugs and toxins can be eliminated from the body through biotransformation processes that occur in the liver.
However, the liver is susceptible to damage from a variety of pharmaceutical agents and environmental chemicals.
Certain medications and their active metabolites have the potential to induce hepatic injury, resulting in conditions such as drug- or toxin-induced hepatitis.
The histopathological architecture of the liver constitutes one of the evaluative parameters for assessing the toxicological impact of various .
A notable substance associated with hepatic injury is diethylene glycol (DEG), a clear and odorless solvent that can occasionally occur as a toxic contaminant in pharmaceutical The threshold for adverse effects is exceeded when an intake of 0.
5 mg per kilogram of body weight is maintained on a daily basis.
The underlying cause of this toxicity may be attributed to the formation of a toxic metabolite, diglycolic acid (DGA).
In rat models, alcohol dehydrogenase (ADH) in the liver metabolizes DEG into the intermediate compound 2hydroxyethoxyacetaldehyde (HEAA) and DGA, which subsequently leads to metabolic acidosis and resultant cellular injury in hepatocytes and the .
Recently, herbals have gained popularity as complementary treatments for inflammation and oxidative stress associated with free radicals and toxic exposure, primarily due to their low toxicity .
One of the notable medicinal plants is celery (Apium graveolen.
, a green plant recognized for its antioxidant and antiinflammatory properties.
This plant encompasses a variety of bioactive compounds, including tannins, saponins, apigenin, luteolin, and kaempferol, which serve as antioxidants.
Its therapeutic effects encompass benefits for hypertension, anti-inflammatory actions, diuretic effects, and the reduction of liver fat .
Previous studies have indicated that celery extract may confer protective effects against liver damage induced by toxic agents.
Specifically, celery extract has been shown to reduce inflammation and enhance total antioxidant capacity (TAC) and glutathione (GSH) levels.
Notably, celery has demonstrated hepatoprotective effects in instances of acetaminophen-induced toxicity.
Emad et al.
reported that pretreatment with a low dose of celery .
mg/kg body weigh.
significantly decreased liver enzyme levels (AST and ALT) and inflammatory markers (TNF- and IL-.
in rats exposed to acetaminophen.
The antioxidant effects of celery on inflammation and arthritis have been investigated in a study by Sukketsiri et .
They employed complete Freund's adjuvant in their animal model, while we used diethylene glycol in our study.
Celery at dosages of 250Ae500 mg/kg BW have demonstrated a beneficial effect against liver oxidative stress by lowering the production of liver free radicals and raising the activity of liver antioxidant enzymes.
The hepatoprotective properties of celery were investigated in two earlier studies and our results confirm them.
In the two previous investigations, complete Freund's adjuvant.
and paracetamol.
were utilized, respectively.
In ours, diethylene glycol was used.
Damage to the liver might result from all of the three aforenamed According to the findings of our study and earlier research, celery may have hepatoprotective effects.
The use of diethylene glycol differentiates our study from previous Our choice of diethylene glycol was based on the fatalities caused by diethylene glycol poisoning in syrup preparations that occurred in Indonesia in 2022.
Our study aimed to evaluate the hepatoprotective effects of celery on DEGinduced histopathological features in rat liver tissue.
METHODS
Research design A post-test only laboratory experimental study with control, conducted from July Ae December 2023 in the Pharmacology and Clinical Pharmacy Laboratory.
Pharmacy Department.
Faculty of Health Sciences.
Jenderal Soedirman University and the Anatomical Pathology Laboratory.
Faculty of Medicine.
Gadjah Mada University.
Materials The tools used in this study included a vacuum pump, an analytical balance (Radwag AS R2.
Polan.
, filter paper, a rotary evaporator (Buchi.
Rotavapor R-3.
Switzerlan.
, glassware, a probe, and syringes for oral administration of turmeric ethanol extract and DEG.
Ekowati H.
Waluyo SS.
Endriastuti NE For histological preparation, the following tools were used: coverglasses, microscope glass slides, tissue embedding cassettes, a rotatory microtome (Shandon AS 325.
United Kingdo.
, styrofoam, histoplast, a paraffin dispenser, and a light microscope (CX23.
Chin.
The materials for histology slide preparation included 10% formalin
hematoxylin-eosin
(HE)
(MayerAos Hematoxylin Solution.
USA), paraffin media, 9% NaCl, and 10% buffered neutral formalin (BNF).
Celery powder was obtained from and has been authenticated by the Testing LaboratoryTraditional Medicine Functional Service Unit.
Tawangmangu.
Central Java.
Other materials included DEG (Merck KGaA.
German.
, 96% ethanol (Merck ETHANOL 96% Reag.
Ph.
Eur.
German.
Tween 80, and standard rat feed.
Experimental animals Male Wistar rats .
months old, weighing 200 gram.
were obtained from the Faculty of Pharmacy.
Muhammadiyah Purwokerto University.
Purwokerto.
Indonesia.
The rats were acclimatized for five days, were given standard pelleted feed and water ad libitum, and maintained on a 12:12-hour light-dark cycle.
The sample size for this study was determined based on the number of experimental repetitions, calculated using FedererAos formula: .
Ou 15, where n = number of samples.
k = number of treatments.
Based on these calculations, each treatment group required a minimum of 5 repetitions, leading to a total of 25 experimental animals for the five groups.
To account for potential unforeseen circumstances, one additional animal was included per group, resulting in a total of 30 Wistar rats.
A post-test only control group design was used for this experimental study.
After the acclimatization period, the rats were divided into five groups, with five rats per group.
Group A .
ontrol grou.
received only food and water.
Group B received DEG orally at 3 g/kg BW twice daily for three days, following six days of food and water administration.
Groups C.
D, and E were treated with DEG .
g/kg BW) and following six days of celery extract administration at doses of 100 mg/kg BW, 200 mg/kg BW, and 400 mg/kg BW, respectively.
The celery extract, dissolved in Tween 80, was administered orally twice daily for six days.
On day 14, all rats were sacrificed, and histopathological damage to the kidneys and liver was assessed using hematoxylin-eosin (HE) Preparation of celery extract A total of 1000 grams of celery powder was extracted using 96% ethanol in a ratio of 1:5 for the first maceration and 1:4 for the 3x24 hours.
Filtration was performed every 24 hours, and the filtrate was concentrated using a rotary evaporator to obtain a solvent-free, thick extract.
Hematoxylin-Eosin (H&E) staining On day 14, all rats were sacrificed via cervical dislocation.
The kidneys and livers were removed and fixed in 10% buffered formalin.
After fixation, tissue samples .
-5 cm slice.
were embedded in paraffin, sectioned, and stained with H&E for histological examination.
The staining procedure was performed by the Anatomical Pathology Laboratory.
Faculty of Medicine.
Gadjah Mada University.
Two trained researchers, under the guidance a pathologist, independently examined the specimens using a binocular microscope (Optilab microscope digital camera syste.
at 400x magnification.
Grading of liver damage Liver damage was scored based on the percentage of injury across four parameters:
Kupffer cell activation, sinusoidal dilation, hepatocyte degeneration, and necrosis.
The scoring criteria were as follows: 0 for normal, 1 for damage <30%, 2 for damage 30-50%, and 3 for damage >50%.
One hundred cells were examined in order to determine the scoring If there is no damage, the score is zero.
there is less than thirty damaged cells, the score is If 30Ae50 cells suffer damage, the score is 2, and if more than 50 cells are damaged, the score is For every organ preparation sample, the results were acquired from five fields of view at a 400x Statistical analysis Liver damage scores were statistically The Shapiro-Wilk test was used to assess normality of data distribution.
We analyzed four sets of data, of which the necrosis data, was not normally distributed.
Therefore, we performed non-parametric analysis using the Kruskal-Wallis test, followed by the Mann-Whitney U test.
This analysis evaluated statistical differences, with a pvalue <0.
05 being considered significant.
Effect of celery in DEG toxicity Ethical clearance The animal experiments were approved by the Ethics committee of the Faculty of Health Sciences.
Jenderal Soedirman University, under ethical approval number 1108/EC/KEPK/V2023.
RESULTS
The liver's color, size, and consistency can all be examined at a macroscopic level.
Figure 1 illustrates that the livers of groups B and C of rats exhibited a color change to pale brown.
Table 1 shows the macroscopic appearance of each group of Wistar rat livers.
The liver color in the normal group (A) was reddish-brown.
Compared to groups B and C, the liver color changed to pale The liver color in groups D and E was reddish-brown, the same as that of group A.
The four groups are all of the same size and consistencyAifour Histopathological observations of the liver (Figure .
identified various forms of tissue damage.
Hepatocyte degeneration, necrosis, sinusoidal dilatation, and Kupffer cell activation are some of the indicators of liver injury that are observed.
Table 2 shows the histopathological damage score based on scoring.
The Shapiro-Wilk test for normality indicated that the data was not fitted to the normal distribution curve .
<0.
In light of the results, it was reasonable to perform non parametric tests, and Kruskal-Wallis test revealed such differences between the groups.
The Mann Whitney test was then carried out to investigate whether pairwise differences exist.
Data in Table 2 shows that Group B (DEG 3.
had significant the highest average Kupffer cell activation, followed in descending order by Groups E (DEG 3g Celery 400 mg/kb BW).
A (Norma.
C (DEG 3 g Celery 100 mg/kg BW), and D (DEG 3g Celery 200 mg/kg BW) .
<0.
Similarly, the greatest average sinusoidal dilation was found in Group B, with the lowest averages in Groups A.
C, and E, respectively.
Hepatocyte degeneration was most prominent in Group B, with the lowest averages observed in Groups D.
E, and A.
Finally, necrosis was most prevalent in Group B, with no signs of necrosis detected in any of the other groups.
Figure 1.
Macroscopic observations of rat livers Table 1.
Results of macroscopic observations of rat liver organs Treatment Group A (Norma.
B (DEG 3 gram.
C (Celery 100 mg/kg BW DEG 3 gram.
D (Celery 200 mg/kg BW DEG 3 gram.
E (Celery 400 mg/kg BW DEG 3 gram.
Liver Morphology Color Size Reddish-brown 4 cm Pale brown 4 cm Pale brown 4 cm Reddish-brown 4 cm Reddish-brown 4 cm Consistency Firm Firm Firm Firm Firm Ekowati H.
Waluyo SS.
Endriastuti NE Figure 2.
Histological evaluation on the liver of control and experimental groups Note: (A) normal.
(B) DEG 3 g.
(C) Celery 100 mg/kg BW DEG 3 g.
(D) Celery 200 mg/kg BW DEG 3 g.
(E) Celery 400 mg/kg BW DEG 3 g.
( ): Kupffer cell activation.
( ): Hepatocyte degeneration.
( ): Sinusoidal ( ): Necrosis.
Hematoxylin-Eosin stain.
Magnification 400x.
Table 2.
Histopathological liver damage based on scoring Treatment groups DEG 3 g
DEG 3 g
DEG 3 g
Celery 100 Celery 200
(B)
mg/kg BW
mg/kg BW
(C)
(D)
DEG 3 g
Celery 400
mg/kg BW
(E)
44 A 0.
40 A 0.
00 A 0.
60 A 0.
68 A 0.
84 A 0.
60 A 0.
92 A 0.
32 A 0.
44 A 0.
16 A 0.
04 A 0.
68 A 0.
40 A 0.
24 A 0.
00 A 0.
88 A 0.
48 A 0.
24 A 0.
12 A 0.
Normal
(A)
Kupffer cell Sinusoidal Hepatocyte Liver necrosis p value <0.
<0.
<0.
<0.
Note : DEG : diethylene glycol.
* : p<0.
@ indicate Kruskal-Wallis test followed by Man Whytney test.
Means with differing superscript within rows were significantly different at the p<0.
DISCUSSION