# Understanding the Role of Deconjugation of Phase II Metabolites in Wastewater: Implications for Wastewater-Based Epidemiology

## TL;DR

This study examines how phase II metabolites in wastewater change during transport, improving understanding of drug and chemical exposure tracking.

## Contribution

The study challenges assumptions about phase II metabolite deconjugation in sewers and provides a framework for better wastewater-based epidemiology.

## Key findings

- N-glucuronides, O-glucuronides, and sulfates increased after deconjugation, showing persistence in sewers.
- Acylglucuronides completely deconjugated in sewers, showing no concentration increase.
- Conjugate stability over 6 months was driven by the parent structure, not the conjugate type.

## Abstract

Metabolism is a critical
bodily function that facilitates
the removal
of toxic chemical buildup within the body. In wastewater-based epidemiology
(WBE), it is crucial to understand the metabolism of biochemical indicators
(BCIs) because metabolites are indicative of consumption (e.g., illicit
drugs, pharmaceuticals) or unintentional exposure (e.g., pesticides,
endocrine disruptors). Phase I metabolites are more widely studied
in WBE due to a combination of factors, including, but not limited
to, stability and analyte cost. Phase II metabolites are often assumed
to deconjugate within the sewer network due to high native concentrations
of enzymes. This work deconstructs this assumption and demonstrates
how the in-sewer stability of phase II metabolites is dependent on
both the parent structure and the conjugate type. In total, 79 BCIs
were assessed and compared to urinary metabolism studies via time-variable
enzymatic deconjugation using two enzymes, β-glucuronidase and
arylsulfatase. The concentrations of free analytes excreted as N-glucuronides,
O-glucuronides, and sulfates increased following deconjugation, reinforcing
the persistence of these BCIs during transport throughout the sewer
network. Conversely, no concentration increase was observed for acylglucuronides,
demonstrating complete in-sewer glucuronide cleavage. In-freezer stability
of conjugates was also assessed over 6 months, where it was observed
that the stability of the parent structure is the driver of stability
rather than the conjugates themselves, indicating minimal enzymatic
activity upon storage. Overall, this paper presents a framework that
can be deployed to gain a more comprehensive understanding of phase
II metabolism and improve the accuracy of WBE workflows as well as
environmental risk assessment approaches.