使用人類糞便及問卷評估經腸胃道暴露之塑膠微粒與早期健康效應

Abstract

塑膠微粒為一新興汙染物，廣泛存在於我們的生活環境中，主要藉由攝食及呼吸進入到人體中，過去動物研究證實了塑膠微粒的高劑量暴露會對生物造成氧化壓力的改變及誘導發炎反應，但仍尚未明確的闡明塑膠微粒對人體的負面影響。近年來也有一些研究在人類糞便檢測到塑膠微粒，證實了塑膠微粒會被我們食入及排出，但目前尚未有人類流行病學研究塑膠微粒暴露評估的標準方法。因此，本研究的目的為：在實驗室中發展一個可行的糞便中塑膠微粒檢測方法，合併使用問卷作為塑膠微粒經腸胃道暴露的評估，並探討塑膠微粒攝入的指標行為及其對氧化壓力及發炎因子的影響。 本研究通過國立臺灣大學行為與社會科學研究倫理委員會 (Institutional Review Board, IRB) 的認可，在大台北都會區招募了五十位20至35歲的健康成年人，請受試者填寫基本資料與塑膠微粒暴露相關習慣的問卷，並記錄6天塑膠食品容器的使用與飲食紀錄，同時也收取糞便、血液、尿液檢體。糞便使用自動顯微紅外光譜法進行塑膠微粒的定性與定量分析。氧化壓力指標使用UPLC-MS/MS分別進行血漿中丙二醛 (Malondialdehyde, MDA) 和尿液中8-OHdG、8-NO2Gua、8-IsoPF2α及HNE-MA的量測。血清樣本使用ELISA分析促發炎因子Interleukin-1α (IL-1α)、Interleukin-1β (IL-1β)、 Interleukin-6 (IL-6)、Interleukin-10 (IL-10)、Tumor Necrosis Factor-α (TNFα)及Interferon- γ (IFNγ) 的表現量。 我們的研究結果表示，50個糞便檢體中，有42個檢測出塑膠微粒，平均每克乾種的糞便中檢測出1699±4233顆塑膠微粒，並辨識出26種不同材質的塑膠微粒，包含Polycaprolactone (PCL) (50%)、Polyurethane (PU) (38%)、Polyvinyl alcohol (PVA) (34%).等，塑膠微粒的形狀依照長寬比分為顆粒 (Pellet)、碎片 (Fragment) 及纖維 (Fiber) 三類，以碎片 (58.7%) 佔最多。然而，兩個空白樣本也分別檢測到1顆及20顆塑膠微粒。 在這個研究中，檢測到了比過去研究中更多種類的塑膠，然而從我們檢測到的塑膠種類來看，塑膠微粒的的來源廣泛且多元，除了塑膠容器以外，包裝 (Packaging) 及生活中其他來源可能在塑膠微粒暴露的貢獻上也扮演了重要的角色。此外，在廣義線性模型中，糞便中塑膠微粒與3天、6天使用塑膠食品容器裝飲品的個數之間有顯著的正向關係 (P < 0.01)，表示使用塑膠食品容器裝飲品的個數對糞便中塑膠微粒的量有一定程度的預測能力。 而問卷及糞便調查塑膠微粒暴露指標則可以看到塑膠食品容器裝飲品的個數、6天FCPs裝熱食個數、3天外食餐數、3天FCPs裝熱食餐數、使用茶包的頻率及咬吸管的頻率與部分氧化壓力指標在統計上有顯著正向關係，可能為塑膠微粒暴露的重要行為。進一步分別使用FCPs裝飲品的個數、是否有咬吸管習慣把受試者分為兩組，可以觀察到FCPs使用量多及有咬吸管習慣的組別在氧化壓力及發炎因子指標的表現量都偏高，但無統計上顯著差異。 總的來說，目前所使用塑膠微粒暴露評估方法還有改善空間，但即便目前的量測方式還沒有辦法準確的估計出塑膠微粒的暴露量，這個研究的結果仍為暴露來源及評估方法帶來了相當多的資訊，且不可否認塑膠微粒對人類的健康負面影響。

Plastic particles are an emerging pollutant that widely exists in our living environment and enter our body through ingestion and inhalation. Previous animal studies have confirmed that high-dose exposure to microplastics and nano-plastics can cause changes in oxidative stress and induce inflammation, but the impact of microplastics on the human has not been clearly clarified. Microplastics (MPs) have been detected in the human fecal samples, suggesting that microplastics can be ingested and excreted by the feces. However, MPs in humans has not been systemically characterized. The data on the dietary behavior contributing to the exposure of MPs are also limited. Therefore, the purposes of this study are to assess the microplastics exposure by using fecal samples in young population, and to identify the indicator from questionnaire for the high level of microplastics exposure on ingestion. Further, I would like to explore the impact of MPs exposure on oxidative stress and pre-inflammatory markers. In this study, we characterized the MPs in the fecal samples, then examined the association of MPs with the dietary behavior obtained from the questionnaire. Lastly, the correlation between the exposure and the oxidative stress and inflammation were tested. Study subjects were recruited from the metropolitan Taipei area. Questionnaires were administered to obtain dietary behavior related with the release of MPs 6 days and 3 months prior to the collection of feces. Fecal samples, peripheral blood and urine were also collected. MPs characterization including the number, components and shapes was determined from the fecal samples using micro-FTIR. Markers of reactive oxygen species (ROS) in urine were tested using UPLC/MS-MS, and markers of proinflammatory cytokines in the peripheral blood were determined using ELISA. A total of 50 subjects with mean age of 24 was included in the final analysis. The mean concentration of MPs was 1699 per gram of feces (dry weight) with the size ranging from 9 to 1334 μm. Twenty-six types of MPs were identified in the feces. Polyimidazoline was present in 52% of the samples, followed by polycaprolactone (PCL) (50%), polyurethane (PU) (38%), and polyvinyl alcohol (PVA) (34%). The identified microplastics were mostly shaped as fragments (58.7%). However, there were also 1 MP and 20 MPs detected in the blank controls, respectively. In this study, more types of plastics were detected than in previous studies, suggesting that the sources of MPs are wide and diverse. In addition to plastic containers, packaging and other supplies in our life may also play an important role in MPs exposure. MPs were correlated with the usage of plastic cups for beverage in 6 days and 3 days (Generalized linear model, p-value < 0.01). Fecal MPs were not correlated with ROS and inflammatory markers. In the microplastics exposure indicators of questionnaire, a statistically significant positive correlation can be observed between the usage of FCPs for beverage and for hot foods, the times of takeaway meals in 3 days, the habit of teabags usage and the habit of straw chewing, and some oxidative stress markers. These behaviors may be important for MPs exposure. Further, we divided the subjects into two groups by using the usage of FCPs for beverages and whether they have the habit of straw chewing. However, the group used more plastic cups for beverage and had straw chewing habit had higher levels of ROS markers of MDA, 8-OHdG, 8-NO2Gua, HNE-MA, 8-isoPGF2α, and inflammatory markers of IL-1α, IL-1β, IL-6, IL-10, IFNγ, and TNFα, although they were not statistically significant. In conclusion, the components of microplastics in fecal samples identified in our study were not confined to the components reported in the previous studies. In addition to plastic containers, plastic food packaging may be an important source for the uptake of MPs. However, this study provided more information for development of microplastics exposure assessments, and it is undeniable that microplastics have some impacts on human health. In the future, we need to develop a better methodology for the exposure assessment of MPs and further identify the sources of MPs.