INTRODUCTION:

Food-contact materials can release chemicals into food, affecting safety and quality. Newspapers are still used as food wrappers in many developing countries, potentially causing chemical contamination due to printing inks containing mineral oils, pigments, solvents, and additives^1–3. Repeated exposure to these chemicals may lead to toxic effects, particularly in individuals consuming food frequently wrapped in newspapers^4–6. Newspaper inks contain pigments, resins, solvents, and additives such as mineral oils, which may migrate into food^7–13. Heavy metals like Cd, Pb, Hg, and Cr, present in ink, pose health risks^14–16.

Polycyclic aromatic hydrocarbons (PAHs) and other compounds may also contaminate water or food sources, affecting humans via the food chain^17,18.

Xylophagia, the compulsive consumption of paper, is a form of pica linked to nutritional deficiencies and neuropsychiatric conditions^19–24. Zebrafish serve as a reliable model for toxicology studies while adhering to the 3Rs principle of reducing animal experiments²⁵.

The present study was conducted to:

1. Evaluate whether newspaper material causes liver toxicity in zebrafish.

2. Raise awareness against using newspapers for food packaging.

3. Investigate pica-related issues in the context of newspaper consumption.

MATERIALS AND METHODS:

Study Site and Duration:

The study was carried out in the Pharmacology Laboratory, RMES College of Pharmacy, during the period from 13 September to 10 October 2022.

Experimental Animals:

A total of 48 adult zebrafish (Danio rerio) were used in the study. Fish were maintained in eight round glass ponds under standard laboratory conditions with a natural light–dark cycle. Animals were fed twice daily with commercially available OPTIMUM aquarium feed and were acclimatized prior to the experiment.

Chemicals:

Newspapers were procured from a local fast-food stall. Aqueous newspaper extracts were prepared using distilled water in graded concentrations ranging from 0.5% to 10%.

Preparation of Newspaper Extract:

Newspapers were collected from a local stall. Aqueous extracts were prepared by soaking weighed newspaper material in distilled water to obtain graded concentrations ranging from 0.5% to 10%.

Experimental Design:

Fish were divided into eight groups (n=6 per group):

Group I: Control

Group II–VIII: Newspaper extract at 0.5%, 1%, 2%, 4%, 6%, 8%, and 10%, respectively.

Experimental Procedure:

Fish were exposed to newspaper extract during morning hours and transferred to clean water in the evening. Treatment was continued for four weeks. Animals were observed daily for behavioral changes and mortality.

Histopathological Evaluation:

At the end of the study, representative liver and intestinal tissues were collected, fixed in 10% neutral buffered formalin, processed by routine paraffin embedding, sectioned at 4–5 µm, and stained with hematoxylin and eosin. Sections were examined under a light microscope.

Semi-quantitative Histopathological Scoring:

Histopathological changes were assessed using a semi-quantitative scoring system for necrosis and inflammation. Multiple non-overlapping microscopic fields from each tissue section were evaluated, and the severity of lesions was graded as follows:

· Necrosis:

0 – Absent, 1 – Mild, 2 – Moderate, 3 – Severe

· Inflammation:

0 – Absent, 1 – Mild, 2 – Moderate, 3 – Severe

Statistical Analysis:

Histopathological scores were expressed as mean ± SEM. Statistical comparison between groups was performed using the Kruskal–Walli’s test followed by Dunn’s multiple comparison test. A value of p < 0.05 was considered statistically significant.

RESULTS:

Histopathological examination of liver and intestinal tissues of zebrafish revealed concentration-dependent pathological alterations following exposure to newspaper extract.

The control group (Group I) showed normal hepatic architecture and intact intestinal mucosa with no evidence of necrosis or inflammatory changes (Figure 1).

Fish exposed to lower concentrations (0.5% and 1%; Groups II and III) exhibited mild fatty changes and minimal inflammatory infiltration in the liver, while the intestinal architecture remained largely preserved (Figure 1).

· At 2% concentration (Group IV), liver sections showed inflammation associated with fatty degeneration, along with damage to the intestinal mucosa (Figure 1).

· Fish exposed to 6% extract (Group VI) demonstrated mild inflammation accompanied by ulceration in both liver and intestinal tissue (Figure 2).

· In group VII (8%), liver sections showed pronounced inflammation and congestion, indicating progressive tissue injury (Figure 3).

· At the highest concentration (10%, Group VIII), severe pathological changes including inflammation, hemorrhage and congestion were observed in both liver and intestine (Figure 2).

Semi-quantitative histopathological scoring demonstrated a progressive increase in necrosis and inflammation scores with increasing concentration of newspaper extract. The mean necrosis and inflammation scores for each group are summarized in Table 1. Comparison with the control group showed that treated groups exhibited statistically significant increases in histopathological scores (p<0.05).

Graphical representation of the semi-quantitative scores further confirmed a dose-dependent trend in necrosis and inflammation across the experimental groups (Figure 3 and 4).

Table 1: Mean Necrosis and inflammation scores

Group	Newspaper extract (%)	Necrosis score (Mean)	Inflammation score (Mean)
I	Control	0	0
II	0.5	1	1
III	1	2	1
IV	2	2	2
V	4	2	3
VI	6	1	2
VII	8	2	3
VIII	10	3	3

Figure: 01

Figure: 02

Figure: 03

Figure: 04

DISCUSSION:

Direct contact of greasy food with newspaper ink can cause chemical migration, leading to liver and intestinal toxicity²⁶. Lead and phthalates in newspaper inks may pose health hazards²⁷. Awareness about food contamination from newspapers remains low. Authorities such as FSSAI advise against using newspapers for food storage and packaging²⁸.

Xylophagia, a form of pica, can lead to ingestion of potentially toxic materials, resulting in severe health consequences^29–31. Zebrafish are useful models for studying such toxic effects while following the 3Rs principle.

CONCLUSION:

The present findings underscore the potential health risks associated with newspaper ink ingestion, particularly in vulnerable populations engaging in xylophagia. Public health interventions, awareness campaigns, and adherence to food safety guidelines are essential to mitigate these risks. Zebrafish models serve as an effective platform for investigating the toxicological impact of chemical contaminants, thereby supporting evidence-based recommendations for food safety and behavioral health.

CONFLICT OF INTEREST:

The Author Declare no Conflict of Interest.

ACKNOWLEDGMENTS:

· RMES College of Pharmacy, Kalaburgi for laboratory facilities

· Asra Histopathology Lab and Dr. Zeenath Begum for histological examination and images

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Received on 17.11.2025 Revised on 02.02.2026

Accepted on 21.03.2026 Published on 22.04.2026

Available online from April 24, 2026

Res.J. Pharmacology and Pharmacodynamics.2026;18(2):127-130.

DOI: 10.52711/2321-5836.2026.00017

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