Zebrafish embryos swimming through a swirling green and brown solution, symbolizing chlorophyll and tannins interference in research.

Natural Product Drug Discovery: Are Hidden Compounds Messing with Your Results?

Jordan Keane in Science & Nature December 2025 • 4 min read.

"Tannins and chlorophylls could be skewing your zebrafish assay results. Learn how to spot and handle these interferences for more reliable natural product research."

Natural products offer a rich source of compounds for drug development. However, their complex composition can create challenges in identifying active molecules using high-throughput screens (HTS). Many assays rely on colorimetric or luminescence-based measurements, which can be disrupted by the presence of tannins and chlorophylls, potentially leading to false positives or negatives.

Zebrafish assays offer an alternative approach to bioprospecting naturally occurring bioactive compounds, yet the interference potential of tannins and chlorophylls in these assays has remained unclear. A recent study investigates the extent to which these compounds can affect the outcomes of zebrafish phenotypic assays, and offers insights into optimizing your research.

This article explores the findings of that study, focusing on how tannins and chlorophylls can interfere with the efficacy of known small-molecule inhibitors in zebrafish embryos. By understanding these interferences, researchers can develop more reliable screening methods for identifying promising drug candidates from natural sources.

Tannins and Chlorophylls: The Usual Suspects in Assay Interference

Zebrafish embryos swimming through a swirling green and brown solution, symbolizing chlorophyll and tannins interference in research.

Tannins and chlorophylls are common secondary metabolites found in plants. Tannins, known for their protein-binding properties, can interfere with enzymatic assays and radioligand receptor-binding assays. Chlorophylls, with their strong light absorption, can disrupt colorimetric assays. Both have the potential to skew results in high-throughput screens, leading to inaccurate identification of active compounds.

While methods exist for removing tannins and chlorophylls from natural product extracts, these processes can be time-consuming and may also remove bioactive compounds. The study by Samat et al. sought to determine whether tannins and chlorophylls could interfere with zebrafish phenotypic assays, which are increasingly used in drug discovery.

Tannins: Can bind to proteins, interfering with enzyme and receptor assays.
Chlorophylls: Absorb light strongly, disrupting colorimetric measurements.
Both: Can lead to false positives or false negatives in high-throughput screens.

The study focused on several key areas to assess this interference. First, tannin-enriched (TEF) and chlorophyll-enriched (CEF) fractions were isolated from Camellia sinensis (tea leaves). These fractions were then tested alone and in combination with known small-molecule inhibitors in zebrafish embryos. The researchers examined various phenotypic endpoints, including body length, trunk curvature, and blood vessel development, to determine if tannins and chlorophylls altered the effects of the inhibitors.

Key Takeaways and Implications for Researchers

The study revealed that tannins and chlorophylls can indeed interfere with zebrafish phenotypic assays, although the extent of interference depends on the specific target and assay. For example, tannin-enriched fractions (TEF) partially masked the antiangiogenic effect of sunitinib malate, while chlorophyll-enriched fractions (CEF) enhanced the activity of γ-secretase inhibitors, leading to toxicity.

These findings underscore the importance of carefully optimizing assays and assessing the interference potential of tannins and chlorophylls before conducting screening assays. Researchers should consider the following:

By understanding and addressing these potential interferences, researchers can improve the reliability of zebrafish assays and increase their chances of identifying promising drug candidates from natural products. While this article showcases this with natural extracts isolated from Camellia sinensis, you can also assess this on different plants and species with similar parameters.

About this Article -

This article was crafted using a human-AI hybrid and collaborative approach. AI assisted our team with initial drafting, research insights, identifying key questions, and image generation. Our human editors guided topic selection, defined the angle, structured the content, ensured factual accuracy and relevance, refined the tone, and conducted thorough editing to deliver helpful, high-quality information.See our About page for more information.

This article is based on research published under:

DOI-LINK: 10.1089/adt.2017.833, Alternate LINK

Title: Interference Potential Of Tannins And Chlorophylls In Zebrafish Phenotypic-Based Assays

Subject: Drug Discovery

Journal: ASSAY and Drug Development Technologies

Publisher: Mary Ann Liebert Inc

Authors: Norazwana Samat, Mei Fong Ng, Nur Faizah Ruslan, Kazuhide Shaun Okuda, Pei Jean Tan, Vyomesh Patel

Published: 2018-10-01

Everything You Need To Know

How do tannins and chlorophylls typically interfere with drug discovery assays?

Tannins, known for their protein-binding properties, can disrupt enzymatic assays and radioligand receptor-binding assays. Chlorophylls, due to their strong light absorption, can interfere with colorimetric assays. This interference can lead to inaccurate identification of active compounds in high-throughput screens, resulting in either false positives or false negatives.

How do tannin-enriched fractions (TEF) and chlorophyll-enriched fractions (CEF) specifically affect zebrafish assays, especially concerning known inhibitors like sunitinib malate and γ-secretase inhibitors?

In zebrafish assays, tannin-enriched fractions (TEF) can mask the effects of certain drugs, such as the antiangiogenic effect of sunitinib malate. Chlorophyll-enriched fractions (CEF) can enhance the activity of other drugs, like γ-secretase inhibitors, potentially leading to increased toxicity. The specific impact of tannins and chlorophylls depends on the target and assay in question.

Why are tannins and chlorophylls problematic in natural product research, and what makes their removal from extracts challenging?

Tannins and chlorophylls are commonly found in plant extracts and can interfere with various assays used in drug discovery. Tannins bind to proteins, disrupting enzyme and receptor assays. Chlorophylls absorb light strongly, which can disrupt colorimetric measurements. Removing them can be challenging because the removal processes are time-consuming and may remove bioactive compounds of interest.

What experimental approach was used to investigate the interference of tannins and chlorophylls in zebrafish phenotypic assays?

The study by Samat et al. used tannin-enriched fractions (TEF) and chlorophyll-enriched fractions (CEF) isolated from Camellia sinensis (tea leaves) to test their effects on zebrafish embryos. Researchers observed endpoints like body length, trunk curvature, and blood vessel development to see if tannins and chlorophylls altered the effects of known small-molecule inhibitors. This approach helped determine the extent to which these compounds can interfere with zebrafish phenotypic assays.

Besides tannins and chlorophylls, what other types of hidden natural compounds might interfere with bioactivity assays, and why?

While the focus is on tannins and chlorophylls, other natural compounds could also interfere with assay results. Compounds with chromophoric properties, like flavonoids or quinones, might affect colorimetric assays. Compounds with detergent-like properties might disrupt cell-based assays. Understanding the chemical properties of the extract and potential interactions with assay components is crucial for reliable results.