Combined Physical and Biological Removal of Antihypertensive Pharmaceuticals from Wastewater: A Case Study from Selected WWTPs ^†

Abstract

Antihypertensive drugs exhibit high resistance and low biodegradability, which leads to their insufficient removal in conventional treatment processes. This study focuses on the removal of selected pharmaceuticals from wastewater using physical–biological methods. These methods included pretreatment by an electrostatic field and biodegradation by a mixed culture of Rhodococcus bacteria.

1. Introduction

Antihypertensive drugs are a group of medications primarily used to treat high blood pressure, which is one of the leading causes of cardiovascular disease [1]. According to the National Health Information Institute [2] in the Czech Republic, the number of patients with hypertension increased by almost 400,000 between 2013 and 2023. This means that one in five Czechs is being treated for high blood pressure.

This trend is also reflected in the consumption of antihypertensive drugs, which has been growing for a long time. With the increasing number of patients taking these drugs, a higher occurrence of active substances and their metabolites in wastewater can also be expected. According to the Organisation for Economic Co-operation and Development (OECD), patients excrete 30–90% of the administered drug dose in the form of metabolites or unchanged form in urine or feces [3,4]. Residues often exhibit high persistence and low biodegradability, leading to their inadequate removal in conventional treatment processes and further release into the environment.

From the group of antihypertensive drugs, diuretics such as furosemide may be prescribed for high blood pressure. It is a loop diuretic that inhibits the reabsorption of Na⁺/K⁺/2Cl⁻ in the ascending part of the Henle loop, thereby increasing water excretion in the urine and lowering blood pressure. Furosemide can be metabolized in the body to glucuronide, saluamine furosemide, or 4-chloro-5-sulfamoylanthranilic acid, but most of it (60% in adults, 84.5% in children) is excreted unchanged [5]. The resulting metabolites can be removed from conventional wastewater treatment plants (WWTPs) by almost 80%. However, the active substance itself is persistent in its unchanged form in conventional treatment processes [6].

Another drug prescribed for high blood pressure is metoprolol, which belongs to the group of beta-blockers for hypertension. These beta-blockers slow down the heart rate and reduce the strength of the heartbeat, thereby helping to lower blood pressure [7]. Metoprolol is one of the most commonly prescribed beta blockers, accounting for over 80% of total consumption in Europe together with atenolol. The specific proportion of individual substances varies, of course, depending on the country and time period [8]. In 2019–2020, metoprolol ranked 9th among the most commonly used drugs in the Czech Republic [9]. Metoprolol is metabolized in the body to alpha-hydroxy-metoprolol and 4’-hydroxy-metoprolol. Approximately 95% of the oral dose is excreted in the urine, with only 5–10% being unchanged; in individuals with slow metabolism (so-called slow metabolizers), this proportion may rise to 30–40% [7,10].

Telmisartan is also a commonly prescribed antihypertensive drug. It is an antihypertensive drug from the group of angiotensin II receptor antagonists (known as sartans), which blocks the binding of angiotensin II to AT₁ receptors, causing vasodilation and a reduction in blood pressure [11]. No active metabolites are formed after ingestion, and the substance is therefore excreted unchanged [11,12].

All of the above-mentioned drugs have also been detected in surface waters in the Czech Republic and have been classified as among the most frequently found drugs in surface waters in the Czech Republic [13,14]. Telmisartan, for example, has been detected in 99% of 256 profiles in surface waters in the Czech Republic over a long period of time, and high concentrations have been measured since 2020 [15].

A large number of studies have investigated ways of removing the above-mentioned drugs. For example, furosemide has been degraded using bacteria such as Agrobacterium tumefaciens and Arthrobacter ureafaciens [16] or fungi such as Aspergillus candidus, Cunninghamella echinulata, and Trametes versicolor [17]. Furthermore, its photodegradation has been described, whereby furosemide reacts to visible light, sunlight, or UV radiation. In the field of innovative processes, the results are often variable. Oxidation methods using hydrogen peroxide and UV radiation show good efficacy. The use of absorption on xylitol, lignite or activated carbon (in powder or granular form) also shows interesting results, with an efficacy of more than 60% [5]. Advanced methods such as the use of electrochemical oxidation techniques have also been investigated for the degradation of metoprolol. Research has shown that the use of conductive diamond electrochemical oxidation can very effectively decompose metoprolol in wastewater [18]. In the field of advanced methods, heterogeneous catalytic ozonation, investigated by Nannou [19], has shown promise, demonstrating effectiveness in removing various pharmaceuticals, including telmisartan, in a continuous flow setting.

All of the above-mentioned pharmaceuticals (metoprolol, furosemide, telmisartan) are persistent in conventional wastewater treatment processes and thus continue to enter the environment. This study focused on the possibility of removing multiple types of pollutants using biodegradation preceded by physical pretreatment. Physical pretreatment uses the advanced oxidation methods mentioned above and consists of exposing the sample to an electrostatic field, assuming the breakdown of the individual matrixes, which facilitates the following biodegradation by a bacterial mixture of Rhodococcus degradans, Rhodococcus erythropolis and Rhodococcus rhodochrous.

2. Materials and Methods

Wastewater samples were taken before entering the recipient from a selected treatment plant in the Moravian-Silesian Region. These were continuously (24-h) combined samples. All glass bottles were purified before sampling. The glass bottles were stored in cooling boxes both during sampling and transport to the laboratory. All samples were sent for initial HPLC/MS/MS analysis to the ALS Czech Republic, s.r.o. laboratory in Ostrava. Some of the samples were first exposed to physical pretreatment on a redesigned patented device for physical waste treatment (EP 2388068) [20], which is presently designed as a laboratory test device. The device can be used for combined or single pre-treatment. Within this experiment, the samples were subjected to an electrostatic field. The technical design of the device is shown in Figure 1. For subsequent biodegradation, bacteria of the genus Rhodococcus from the Czech Collection of Microorganisms at the Faculty of Science, Masaryk University in Brno were used.

Figure 1. Technical design of the device for physical waste pre-treatment. Legend: 1, 2, 3, 4, 5—generators of force fields (spark discharge–high temperature plasma, microwave field, ultrasound, UV radiation, electrostatic field–non-thermal plasma), 6—plastic sample container, 7—sample ready to testing, 8—Faraday cage, 9—supporting bridge carrying generators 1–5, including their drives, switches, and circuit breakers, 10, 11—conductive metal grid, 12 a, b—HV supply cables connected to generator 5 (electrostatic field) (Reprinted from Refs. [20,21]).

• Laboratory Analysis

• Procedure for Preparing Wastewater Testing3

The wastewater samples were submitted for primary HPLC/MS/MS analysis to the ALS Czech Republic, s.r.o. laboratory in Ostrava. Afterwards, some of the samples were submitted for physical pretreatment to T. G. Masaryk Water Research Institute (VUV TGM) in Ostrava, where the samples were subjected to physical pretreatment on an EP 2388068 device under the effect of an electrostatic field at a voltage of 15 kV and a current of 0.1 mA for 8 h.

After physical pretreatment and transport of the samples to the laboratory, 1 L water samples were poured into 2 L biodegradation beakers using the following procedure:

“Bacteria” samples

A combination of 1 L of water (without physical pretreatment), 200 mL of M011 Soyabean Casein Digest Medium (Tryptone Soya Broth) culture medium, and 200 mL of a mixed culture of Rhodococcus degradans (4446 CCM), Rhodococcus erythropolis (277 CCM), and Rhodococcus rhodochrous (2751 CCM).

Samples “physical + bacteria”

A combination of 1 L of water after physical pretreatment, 200 mL of culture medium M011 Soyabean Casein Digest Medium (Tryptone Soya Broth) and 200 mL of a mixed culture of bacteria Rhodococcus degradans (4446 CCM), Rhodococcus erythropolis (277 CCM) and Rhodococcus rhodochrous (2751 CCM).

Control samples

A combination of 1 L of water (not physically pretreated) and 400 mL of distilled water.

“Physical control” samples

A combination of 1 L of water after physical pretreatment and 400 mL of distilled water.

All measurements were carried out at a temperature of 23 °C, pH 8.1, under continuous oxygenation using JK-AP7500 360 L/h aerators (JK Animals, Chotěšov, Czech Republic). The samples were biodegraded over a period of 20 days. During the measurements, distilled water was regularly added to prevent evaporation of the liquid phase.

• Sample Analysis

All measurements were finished after 20 days. The samples were then placed in pre-purified glass bottles and transported in cool boxes to the ALS Czech Republic, s.r.o. laboratory in Ostrava for HPLC/MS/MS analysis.

3. Results and Discussion

We will examine the results for metoprolol in more detail, as the data for furosemide and telmisartan are inconclusive. This is consistent with studies by Olvera-Vargas, Liu and Kaiser [22,23,24], which state that these two drugs are not readily biodegradable. However, we can at least compare the values of the drug concentrations we detected in the treatment plant with other studies. In Europe, the median concentration of furosemide in the influent to WWTPs is around 2.6 μg/L, ranging from 0.525 μg/L for Iceland to 13 μg/L for the Czech Republic. The average concentration of furosemide is around 4.4 μg/L, and the average concentrations are relatively close to the medians, with the exception of the Czech Republic, where Rozman [25] found a maximum concentration of 71.5 μg/L for the municipality of Onšov [5]. Our measured data of 0.814 μg/L and 1.221 μg/L are among the lower values in this range. Telmisartan was detected in wastewater at concentrations of 0.69 μg/L [26], 4.35 μg/L [27], and 2.3 μg/L [28]. Our measured data on telmisartan concentrations of 1.093 μg/L and 0.101 μg/L are again among the lower values.

The metoprolol concentrations we measured, 0.693 μg/L and 0.401 μg/L, correspond to the range reported in studies by Maurer and Scheurer [10,29], which report detected concentrations in the range of 0.160–2 μg/L. Figure 2 and Figure 3 below show and compare the metoprolol concentrations we measured in 2023 and 2024.

Figure 2. Measured concentrations of metoprolol from samples taken in 2023.

Figure 3. Measured concentrations of metoprolol from samples taken in 2024.

Metoprolol responded well to biodegradation by Rhodococcus bacteria. In the 2023 sampling, biodegradation reduced the concentration of metoprolol in the samples by 70%. Using a combination of physical pretreatment and biodegradation, the concentration in the samples was reduced by 69%, which is a negligible difference.

In 2024, biodegradation reduced the concentration of metoprolol by 94% and physical pretreatment by 95%.

4. Conclusions

Antihypertensive drugs are drugs that should be given increased attention, as their use in the population is growing, and thus, the concentration of these substances in the environment is increasing. Our experiment showed that biodegradation is not very suitable for the groups of diuretics and sartans, but β-blockers respond very well to biodegradation. In the future, the experiment should also focus on shorter biodegradation times to bring the results closer to real conditions in wastewater treatment plants, as well as on new possibilities for combining physical pretreatment with biodegradation to increase the efficiency of combined processes.