Measurement of Microplastic Release After the Use of Polypropylene Nasal Irrigation Bottles
Article information
Abstract
Objectives.
Microplastics originating from plastic materials may pose risks to human health. This study investigated the presence of microplastics in nasal irrigation fluids collected from reused bottles, focusing on the duration of bottle usage.
Methods.
Readily available nasal irrigation bottles made of polypropylene were purchased. Unused bottles served as controls. Test samples were prepared to simulate 1-, 3-, and 6-month reuse. Nasal irrigation fluid samples (n=12) were collected from each set of bottles: three from the new control bottles and nine from the bottles simulating 1-, 3-, and 6-month reuse. Raman spectroscopy was used to detect microplastics in the nasal irrigation samples, and the results were compared based on the duration of bottle use.
Results.
An average of 33.00±20.42 microplastic particles per 300 mL was detected in the nasal irrigation fluid from the control bottles. In comparison, bottles used for 1, 3, and 6 months contained averages of 68.66±30.07, 261.66±20.59, and 204.33±52.16 microplastic particles per 300 mL, respectively. The majority of these particles ranged in size from 10 to 100 μm and were primarily fragment-shaped. Polypropylene was identified as the predominant type of microplastic, suggesting it was directly released from the irrigation bottles.
Conclusion.
We detected microplastics in nasal irrigation fluids, which likely originated from the repeated use of nasal irrigation bottles. The quantity of microplastics was significantly higher in samples from bottles simulating 3 months of use compared to the control samples. Therefore, we recommend the development of guidelines to regulate the duration of nasal irrigation bottle usage to reduce microplastic infiltration into the body via the sinonasal cavity.
INTRODUCTION
Plastic particles sized <5 mm are referred to as microplastics (MPs) [1]. These particles originate either from the degradation of larger plastic pieces or are deliberately produced for use in various industries [1]. MPs are pervasive in the environment, appearing in soil, seas, food, the atmosphere, and engineered systems [2-4]. The widespread presence of MPs poses significant risks to both human health and ecological systems. For example, when released into aquatic environments, MPs can be ingested by marine organisms and subsequently accumulate in the food chain. Additionally, MPs in the atmosphere may contribute to climate change and lead to respiratory diseases [5-7].
Inhalation, ingestion, and dermal exposure are the three primary pathways through which MPs enter the human body [8]. In 2019, MPs were detected in human feces, indicating inadvertent ingestion from various sources [9]. Previously, the presence of MPs has been documented in the human placenta, including the maternal, fetal, and amniochorionic membranes [10]. Amato-Lourenco et al. [5] also identified MPs in the lung tissues of non-smokers. Furthermore, multiple types of MPs have been found in venous blood samples from healthy, non-fasting adult volunteers [11].
The nasal cavity often serves as the first line of defense against pathogens entering the human respiratory system by trapping these invaders. Consequently, the presence of MPs in the sinonasal cavity and their effects are significant areas of interest. MPs that enter the nasal cavity can directly harm nasal tissues, potentially facilitating their transport to other organs through nasal penetration [12]. Additionally, it has been reported that MPs may cause pulmonary toxicity through various mechanisms, including inflammasome activation, DNA damage, and mitochondrial injury [13-15]. Despite these concerns, few studies have explored the presence of MPs in the sinonasal cavity. The detection of MPs has been documented in nasal lavage fluids from healthy volunteers, as well as patients with allergic rhinitis and chronic rhinosinusitis [1,16]. Given that these studies involved the collection and analysis of nasal lavage fluid samples, we hypothesized that not only airborne MPs but also nasal irrigation bottles could be sources of MP contamination. Notably, the recycling of plastic bottles has been shown to facilitate the release of MPs [17]. Since nasal irrigation bottles are typically made from plastic and reused multiple times, we hypothesized that nasal irrigation fluids from these bottles might also be a significant source of MP release.
In a preliminary study, we suggested that MPs might be present in nasal irrigation fluids [18]. To further confirm this hypothesis, we assessed the presence of MPs in nasal irrigation fluids collected from nasal irrigation bottles, categorizing them according to their duration of use.
MATERIALS AND METHODS
Preparation of samples
Readily available nasal irrigation bottles made of polypropylene (PP) with Europe’s CE (Conformité Européenne) mark were purchased (Nasalfresh, Pinpointercommunications). Unused irrigation bottles served as the controls. Nasal irrigation bottles mimicking 1-, 3-, and 6-month reuse were prepared as test samples. Each sample group comprised 3 bottles, and 12 nasal irrigation bottles (3 for the control samples and 9 for the reused samples) were employed in the analysis. Reused nasal irrigation bottle samples were prepared by cleaning them 30, 90, and 180 times (once daily) to mimic 1, 3, and 6 months usage, respectively, following the manufacturer’s protocol. The manufacturer’s protocol instructed that the purified water-filled bottle be heated in a household microwave for 30 seconds to reduce the risk of contamination and infection. The cleaning tip and connecting tube were boiled in hot purified water for 30 seconds. The protocol for harvesting irrigation fluids from each nasal irrigation bottle for MP analysis was as follows: approximately 300 mL of ultrapure water was added to the prepared bottles, and the nozzle cap was closed tightly. The samples were then vigorously shaken at least 10 times, and the fluids were collected. The obtained fluid samples were filtered through a 5-μm Silicon filter (10×10 mm2, Smart membrane) for the collection of MPs.
Raman analysis
The Raman analysis procedures were performed following a previous protocol using an XploRA Plus confocal Raman microscope [18]; a 532-nm laser and a 1,024×256 pixel-cooled charge-coupled device detector were employed for the Raman analysis. The laser power was decreased by 10% by using an embedded filter. Gratings with 1,200 grooves/mm were employed in this study. The confocal hole and slit width were set to 100 μm and 50 μm, respectively. A system calibration was performed by zero-order correction of the grating and additionally on the 520.7 cm–1 peak of silicon. Mosaic images of the microscopic dark-field images of the filter surface were obtained using an M Plan Semi Apochromat BD objective lens 20×/N. A. 0.45 (Olympus), and the images were processed using the ParticleFinder module in LabSpec 6 software. All bright particles obtained using this procedure were selected from the dark background area for analysis. Raman spectra were recorded by accumulating a 1×2-second exposure time in the spectral range of 1,020–3,400 cm–1. LabSpec 6 software was employed to process the collected spectra. A polynomial baseline was employed for baseline correction, and spectra with high fluorescence were excluded. All spectra were screened for plastics using the classical least-squares algorithm (CLS) (Supplementary Fig. 1). Polyethylene (PE), PP, polystyrene (PS), polyvinylchloride, polyurethane, polyacrylonitrile-butadiene-styrene, polyamide, alkyl acrylate copolymer, and polyethylene terephthalate (PET) spectra were used as standard references for CLS fitting. Each measured spectrum was derived from the summation of all references and theoretical compositions. The CLS spectrum results were manually examined to prevent any potential data omissions or false positives caused by the comprehensive spectrum matching software that utilizes a Raman library.
Quality control
To mitigate the risk of MP contamination, plastic tools were omitted, and glass materials were exclusively utilized throughout the sampling and filtering procedures. All the procedures, including sample preparation and filtration, were performed inside a laminar flow box (SINAN Science Industry, HSCV-1300) to exclude the possibility of contamination from indoor airborne MPs. All solutions, such as ultrapure water and chemical reagents, were previously filtered using glass fiber filter/grade F (medium, high loading grade) and metal filters (0.5 μm) before use. All glassware was cleaned with filtered ultrapure water prior to the laboratory experiments. Samples were wrapped in aluminum foil when transferred outside the laminar flow hood. To reduce the risk of sample contamination, nitrile gloves and cotton coats were donned during processing. Three blank samples, created using empty glass beakers, were subjected to identical procedures. Their outcomes were compared with those of the experimental groups to assess the impact of potential contamination during sample pretreatment and analysis.
Statistical analysis
All descriptive data are presented as the mean±standard deviation. Intergroup differences were evaluated using one-way analysis of variance, followed by post-hoc analysis. All statistical analyses were performed using GraphPad Prism version 8.0, and P-values <0.05 indicated statistical significance.
RESULTS
MP levels in irrigation fluid samples according to bottle-usage duration
Twelve nasal irrigation fluid samples were collected from 12 nasal irrigation bottles, comprising three control and nine reused bottles. The number of MPs identified by Raman analysis is detailed in Table 1. In the control samples, 99 MPs were detected, while the 1-, 3-, and 6-month-used samples contained 206, 785, and 613 MPs, respectively. No MP particles were detected in three laboratory blank samples (data not shown). The average number of MPs per irrigation sample was calculated as follows: 33.00±20.42 (per 300 mL) for the control, 68.66±30.07 (per 300 mL) for the 1-month-used samples, 261.66±20.59 (per 300 mL) for the 3-month-used samples, and 204.33±52.16 (per 300 mL) for the 6-month-used samples. The differences between groups were statistically significant, with P-values as follows: control versus 3 months (P<0.001), control versus 6 months (P<0.001), 1 month versus 3 months (P<0.001), and 1 month versus 6 months (P<0.05) (Fig. 1).
Number of microplastics according to size range within the control and used irrigation samples
Number of microplastics detected in nasal irrigation fluid samples. *P<0.05, **P<0.01.
Characterization of the MPs in the irrigation fluid samples
The detected MPs were characterized based on their shape, size, and type of polymer, as detailed in Supplementary Table 1. In terms of shape, all MPs were identified either as fragments or fibers, with fragments being the predominant type (Fig. 2). Specifically, the control samples contained 77 fragments (78%) and 22 fibers (22%). In the 1-month-used samples, there were 177 fragments (86%) and 29 fibers (14%); the 3-month-used samples had 699 fragments (89%) and 86 fibers (11%); and the 6-month-used samples included 532 fragments (87%) and 81 fibers (13%) (Fig. 2). Regarding size, MPs measuring 20–50 μm were most commonly found, followed by those measuring 10–20 μm (Table 1). MPs in the size ranges of 5–10 μm and those larger than 100 μm were observed less frequently.
Shape of the microplastics detected in nasal irrigation fluid samples.
The types of polymers detected in the control and reused samples were characterized, identifying four types: PP, PE, PS, and PET (Fig. 3). The control samples contained 96 PP, 2 PE, and 1 PET MPs. After 1 month of use, the samples contained 190 PP, 8 PE, 2 PS, and 6 PET MPs. After 3 months, the samples contained 776 PP, 6 PE, 0 PS, and 3 PET MPs. After 6 months, the samples contained 607 PP, 1 PE, 0 PS, and 5 PET MPs (Supplementary Table 1).
Polymer types of microplastics detected in nasal irrigation fluid samples. PP, polypropylene; PE, polyethylene; PS, polystyrene; PET, polyethylene terephthalate.
DISCUSSION
Our study primarily aimed to determine whether nasal irrigation bottles could be a source of MPs. We discovered that MPs were present in the nasal irrigation fluid samples, which were prepared by simulating actual patient use and cleaning methods. Since the polymer type of the MPs identified in the irrigation fluids was identical to that of the PP bottles, we speculate that the MPs originated from the bottles themselves. Our findings indicated that the number of MPs in nasal irrigation fluid samples from bottles used for 3 months was significantly higher than that in control samples from new nasal irrigation bottles. However, there was no significant difference in the number of MPs between samples from bottles used for 3 months and those used for 6 months. We could not determine the exact reason for the slight decrease, rather than an increase, in the amount of MPs released after 3 months of bottle usage. Further experimental results over a longer period are necessary to draw a definitive conclusion about MP release during extended usage.
Nasal irrigation improves nasal mucosal function through multiple physiological mechanisms, such as directly cleansing the mucus to inhibit bacterial growth, and is recommended for a variety of conditions. It serves as the primary treatment for chronic rhinosinusitis, helping to reduce the overuse of antibiotics. After endoscopic sinus surgery, sinus irrigation is vital for improving the healing of nasal cavity and sinus wounds during the postoperative period, thus diminishing the reliance on unnecessary medications [19]. Given the widespread use of nasal irrigation in rhinology, we recommend the development of guidelines for measuring and managing MPs in nasal irrigation fluids.
MPs have been detected in the air, and these airborne particles can enter the human sinonasal cavity [20]. In China, the concentration of MPs in atmospheric dust has been measured at 31±8 to 43±4 particles/m2/day, with an average deposition rate of 132 particles/m2/day for plastic rain [21]. This results in an annual deposition of over 1,000 tons of plastic onto protected lands in the western United States [22]. Individuals are estimated to inhale between 26 and 130 MPs daily [23]. In a specific case, a mildly active man using a breathing thermal manikin was estimated to inhale 272 indoor airborne MPs per day [24]. Two recent studies have assessed the concentration of MPs in human nasal lavage samples. Tuna et al. found MPs at a concentration of 2.38±1.85 pieces/mL, while Tas et al. [16] reported a concentration of 2.34±1.89 pieces/mL in the nasal lavage fluid of healthy individuals [1]. In our study, we detected MP concentrations of 33.00±20.42, 261.66±20.59, and 204.33±52.16 per 300 mL irrigation fluid in the control, 3-month-used, and 6-month-used samples, respectively. Although the absolute concentrations may not seem high, the significance of MPs in nasal irrigation fluids should not be underestimated, given the repeated and prolonged use of large volumes of these fluids.
Regarding the characteristics of the MPs identified in this study, most were fragment-shaped and ranged in size from 10 to 100 μm. These findings align with our previous preliminary results [18]. To our knowledge, no other studies have explored the characteristics and clinical implications of MPs in the human sinonasal cavity. The characteristics of MPs in the lower airways, however, have shown varied results. For instance, particle widths in the bronchoalveolar lavage fluid samples from adults in Northern Europe ranged from 20 to 283 μm, with an average of 49 μm, and fragments constituted 84.42% of the total MPs [25]. In younger adults in Western Europe, the average size of MPs in bronchoalveolar lavage fluid samples was 1,730±150 μm, and most were microfibers (97.06%) [26]. In Asian children, the majority of MPs found in bronchoalveolar lavage fluid samples were smaller than 20 μm [27]. The characteristics of MPs in the human airway vary widely, and their clinical implications remain underexplored. Therefore, while it may not be feasible to establish detailed guidelines for MP regulation immediately, it is crucial to develop measurement guidelines for the total amount of MPs generated from the reuse of commercially available irrigation bottles as soon as possible.
MPs entering the sinonasal cavity present various health risks through different mechanisms. In vitro studies have shown that MPs cause cytotoxicity in a concentration-dependent manner. Additionally, there is evidence suggesting that MPs can penetrate the nuclei of human nasal epithelium cells, potentially leading to DNA damage [28]. Repeated intranasal application of MPs in mice has been linked to behavioral changes, including decreased food and water intake, and has caused pathological alterations in the nasal mucosa, such as mucosal thinning [28]. MPs entering the nasal cavity have also been found to disrupt the nasal microbial balance, with bacteria such as Staphylococcus frequently associated with MPs in these experiments [29]. Furthermore, MPs inhaled through the nasal airway can travel to the lower airway via nasal penetration and may be absorbed into various human organs [28,30]. However, compared to studies focused on the lower airway, research on the clinical significance of sinonasal MPs is very scarce. Future research should focus on the in vivo effects of various types and concentrations of MPs.
We hypothesized that a small amount of MPs might be released into the irrigation fluid due to minor wear and tear from repeated use of the nasal irrigation bottle. Supporting this, a previous study reported that recycling plastic products can also lead to the release of MPs [17]. Additionally, in our study, the irrigation bottle was subjected to microwave heating and boiling as part of its disinfection process, which could potentially lead to further release of MPs [18]. To accurately pinpoint the sources of MP release, experimental studies that consider various product types and disinfection methods are essential. Our findings indicate that MPs can be detected in fluid from nasal irrigation bottles, and that the quantity of MPs increases with the duration of the bottle’s use. Based on these observations, we recommend the development of guidelines regarding the usage duration of nasal irrigation bottles to prevent MPs from entering the human nasal cavity. However, the impact of MPs on human health via the nasal cavity remains unclear, necessitating further research to explore this issue.
This study has certain limitations. First, our results are based solely on one type of nasal irrigation bottle, and the statistical analysis employed was too simplistic to yield clinically significant results. To broaden the applicability of our results, various nasal irrigation tools must be tested. Second, we did not assess the presence of MPs in human nasal mucosa or their effects in vivo. Conducting studies on nasal irrigation samples collected from individuals before and after nasal irrigation could provide additional support for our hypothesis. Furthermore, research into the clinical implications of MPs in the sinonasal cavity is warranted. Third, the smallest particle size detectable with our current method was 5 µm, and we only identified MPs larger than 5 µm. Recent studies have reported the presence and toxic effects of nanoplastic particles, which are significantly smaller than those detected in our study, in the lower airway [25,31]. Therefore, along with MPs, nanoplastics in the upper airway should be further evaluated.
In conclusion, we demonstrated the presence of MPs in nasal irrigation fluid samples, likely originating from the nasal irrigation bottles themselves. Furthermore, we observed a significant increase in the number of MPs in the fluids from bottles simulating 3 months of use compared to the control samples from new bottles. It is necessary to develop guidelines on the duration of nasal irrigation bottle usage to minimize the infiltration of MPs into the human body through the sinonasal cavity. Further investigations that measure changes in MP concentrations in the human sinonasal cavity before and after irrigation would provide additional support for our findings.
HIGHLIGHTS
▪ Microplastics were observed in nasal irrigation fluids.
▪ The number of microplastic particles significantly increased after longer usage.
▪ Microplastics were 10–100 μm in size, fragment-shaped, and predominantly made up of polypropylene.
▪ Guidelines regulating the duration of nasal irrigation bottle usage are warranted.
Notes
No potential conflict of interest relevant to this article.
AUTHOR CONTRIBUTIONS
Conceptualization: HJM. Methodology: all authors. Visualization: HJM. Funding: HJM. Writing–original draft: HJM. Writing–review & editing: KSK. All authors read and agreed to the published version of the manuscript.
Acknowledgements
This work was supported by a National Research Foundation of Korea (NRF) grant funded by the Korean government (MSIT) (2022R1F1A1063720).
SUPPLEMENTARY MATERIALS
Supplementary materials can be found online at https://doi.org/10.21053/ceo.2024.00182.
Examples of microphotographs and results of the Raman analysis of major microplastic types.
Raman analysis results of the samples