A novel whole-greenhouse static chamber method for quantifying ecosystem respiration and nitrous oxide emissions in greenhouse cultivation systems

Abstract

We present a novel and cost-effective whole-greenhouse static chamber method for quantifying nighttime ecosystem respiration (carbon dioxide emissions in the dark) and nitrous oxide (N₂O) emissions in greenhouse cultivation systems. This method leverages routine nighttime sealing practices—such as closing vents and deploying thermal quilts—to create an enclosed environment for gas accumulation. Gas concentrations can be monitored either automatically using in situ sensors or manually via gas-tight sampling bags. In both approaches, linear regression of gas concentration versus time yields robust accumulation rates, which are then converted into fluxes based on greenhouse volume and temperature. Validation using a high-precision Picarro analyzer demonstrated strong linearity (R² > 0.95) in carbon dioxide (CO₂) and N₂O accumulation curves across 15 greenhouses, confirming the method’s reliability. As both approaches produce comparable results, we recommend manual sampling for its simplicity and cost-efficiency. The method integrates emissions from soil, plants, and other surfaces, overcoming spatial limitations of small chambers. With minimal equipment and labor requirements—especially when using manual sampling—it offers a practical solution for routine monitoring of greenhouse gas (GHG) emissions in controlled-environment agriculture.

• •Allows for whole-ecosystem CO₂ and N₂O flux estimation using manual sampling and gas chromatography.
• •Demonstrates strong agreement with continuous online measurements in multi-site validation.
• •Provides a low-cost, scalable solution for GHG quantification in protected cropping systems.

Graphical abstract

Specifications table

Subject area	Environmental Science
More specific subject area	Greenhouse-scale GHG emission quantification
Name of your method	Whole-greenhouse static chamber method
Name and reference of original method	Su et al. (2021) An automated and continuous static chamber system for greenhouse gas measurement based on soil surface monitoring
Resource availability	Gas-tight sampling bags and access to laboratory gas chromatography for analysis. Optional validation equipment: a multi-component gas analyzer (G2508, Picarro, USA), a gas chromatograph (GC-2014, Shimadzu, Japan), and a multi-channel chamber control system (SF-3000, Lica United Inc., Beijing), standard gas cylinders; greenhouse quilts

Background

Greenhouse cultivation has expanded rapidly worldwide in response to growing demands for high-yield, year-round crop production [[1], [2]]. This intensive form of agriculture often relies heavily on synthetic fertilizers, organic amendments, and controlled environmental conditions, all of which can significantly influence greenhouse gas (GHG) emissions—particularly carbon dioxide (CO₂) and nitrous oxide (N₂O) [[3], [4], [5]]. Despite their importance, accurate quantification of these emissions within greenhouse systems remains challenging due to several technical and methodological limitations [6].

One major constraint is the spatial heterogeneity of soil GHG fluxes. Traditional small static chambers, which are commonly used for field-scale flux measurements, sample only limited soil surface areas and are prone to high variability caused by micro-site conditions, plant root activity, fertilizer hotspots, and inconsistent chamber placement [[7], [8], [9]]. This can result in unreliable extrapolation to the scale of the entire greenhouse. Moreover, small chambers do not capture emissions from non-soil sources, such as plant respiration or microbial processes in mulch layers and irrigation channels, which can be substantial in controlled environments [[10], [11], [12], [13]].

Micrometeorological techniques, such as eddy covariance or flux gradient methods, require open or homogeneous terrain and are thus poorly suited for use in semi-enclosed greenhouse environments with intermittent ventilation, complex airflow patterns, and physical obstructions like support structures and plastic films.

To address these challenges, we propose a whole-greenhouse static chamber approach that leverages the routine nighttime sealing of greenhouses using thermal quilts. By treating the sealed greenhouse as a single large chamber, and tracking linear accumulation of CO₂ and N₂O overnight, this method captures integrated emissions from all ecosystem components—including soil, plants, and other surfaces—without the need for complex instrumentation. This approach enables robust, ecosystem-scale GHG flux estimation under realistic production conditions, offering a scalable and practical solution for monitoring emissions in protected agriculture systems.

Method details

• 1.Greenhouse selection: Inspect each greenhouse structure to confirm that all sealing and insulation components (e.g., quilts, vents) operate correctly without air leaks. Only greenhouses that can be fully sealed are suitable for this method.
• 2.Estimating greenhouse volume and planting area: Measure the greenhouse length (L), width (W), and average internal height (H). Calculate the internal volume (V) as V = L × W × H. The planting area (A) is simply L × W. If measuring both V and A is difficult, one may derive H directly, which is equivalent to the mean height and sufficient for flux calculations (H = V/A).
• 3.Gas sampling for CO₂ and N₂O analysis: After sealing the greenhouse in the evening (e.g., closing the vent and rolling down the quilt), collect gas samples at the center of the greenhouse at multiple timepoints (see Fig. 1). Two options are available: 1) Automated online monitoring: Use a continuous analyzer (e.g., Picarro G2508) with an inlet inside the greenhouse for high-frequency measurements. 2) Manual bag sampling: Draw samples into gas-tight bags and analyze them later in the lab by gas chromatography. We recommend manual air-bag sampling for routine applications because it is simple, rapid, low‑cost, and enables simultaneous sampling across multiple greenhouses. The sampling bags were made of an aluminum-plastic composite film, which has a very low gas adsorption capacity and thus does not affect measurement results. The typical bag volumes are 300 mL or 500 ml. Gas samples should be analyzed within one month, preferably within one week. Gas chromatography was calibrated weekly, and standard gases were injected during every run to ensure accuracy.
• 4.Timing of sampling intervals: Determine the diurnal soil temperature cycle inside the greenhouse. In our study, soil temperature peaked at 15:00; therefore, we sampled between 18:00 and 24:00 (or during a 1-hour representative window such as 20:00–21:00) to capture average daily fluxes. For other locations, shift the sampling window to follow the local soil temperature peak (e.g., delay sampling by 1 hour if the peak occurs at 16:00).
• 5.Calculating accumulation rates: For each sealed period, apply a linear regression of gas concentration versus time to obtain the slope (ΔC/Δt in ppm per day). Convert this to a flux (e.g., kg C ha⁻¹ d⁻¹ for CO₂ or g N ha⁻¹ d⁻¹ for N₂O) using the equation: Flux = (ΔC/Δt) × (V/A) × (M/Vm) × 10 × 273/(273+T). Where, V/A is the volume-to-area ratio (or mean height, m), M is the atomic mass of C (12 g mol⁻¹) or N (28 g mol⁻¹) in 1 mol of CO₂ and N₂O, T is the temperature ( °C) in the chamber during sampling, and Vm is the molar volume at ambient temperature and pressure (m³ mol⁻¹). The result represents the whole-greenhouse ecosystem respiration or N₂O emission rate.

Fig. 1.

Two methods were adopted to observe the changes in the concentrations of CO₂ and N₂O gases in the greenhouse during nighttime. To demonstrate the on-site operation, the photo was taken during the daytime.

Method validation

• 1.Greenhouse-scale accumulation measurements: In Weifang, China, we used a Picarro G2508 to record CO₂ and N₂O concentrations in a greenhouse continuously under sealed, dark conditions, deriving accumulation rates via linear regression. To test reproducibility, we then measured 14 additional nearby greenhouses using the same Picarro setup, confirming method transferability across sites.
• 2.Reproducibility and linearity: Across 15 greenhouses in Weifang, Shandong, nighttime concentration regressions exhibited high linearity (R² > 0.95), demonstrating consistent CO₂ and N₂O accumulation trends under sealed conditions (Fig. 2, Fig. 3). Variability in absolute accumulation rates likely reflects differences in greenhouse age, crop type, fertilization regime, and management practices. Gas concentrations in the greenhouses accumulated linearly, with no observable saturation effect up to ∼1700 ppm for CO₂ and ∼3.88 ppm for N₂O. Any slight curvature in the concentration-time curves was primarily attributable to the overnight temperature drop.
• 3.Temperature effects: Concentration–time curves for both gases exhibited slight downward curvature as greenhouse temperature declined overnight (Fig. 2, Fig. 3). Fig. 4 illustrates the influence of temperature on accumulation rates; modeling confirmed that sampling between 18:00–24:00 (or 20:00–21:00) best represents the daily average flux in this region. Because local time zones and diurnal rhythms differ across regions, the recommended 18:00–24:00 sampling window should be applied flexibly. In practice, it corresponds to approximately 3–9 h after the daily soil temperature peak, provided that the greenhouse is sealed and dark. Users in other regions should adjust the sampling window based on the timing of the local soil temperature maximum.
• 4.Adsorption and dissolution assessment: We evaluated CO₂ and N₂O partitioning into condensate and adsorption onto internal surfaces using conservative assumptions. Even under generous estimates, these processes accounted for < 0.1 % of the total gas present, indicating negligible impact on accumulation measurements (see Supplementary data of Quan et al., 2025 [13]).
• 5.Collectively, these validations confirm that simple manual bag sampling under sealed, dark conditions can accurately quantify whole‐greenhouse CO₂ and N₂O fluxes.

Fig. 2.

Examples showing the gradual linear accumulation of CO₂ in five vegetable greenhouses at night (red line) and the linear fitting relationship based on data from 18:00 to 24:00 (bold black fitting line).

Fig. 3.

Examples showing the gradual linear accumulation of N₂O in five vegetable greenhouses at night (green line) and the linear fitting relationship based on data from 18:00 to 24:00 (bold black fitting line).

Fig. 4.

Diurnal variations in greenhouse-scale ecosystem respiration and N₂O emissions in a greenhouse in Weifang, China, during a 70-day observation period.

Limitations

• 1.Approximate volume estimation: Accurate measurement of greenhouse volume or mean height is difficult, and approximations introduce uncertainty into flux calculations. However, errors are generally within 20 % and have only limited effects on results.
• 2.Requirement for sealed, dark conditions: The method requires full quilt closure and darkness, restricting daytime application. Daytime emissions were estimated using temperature—the dominant driver of diurnal variation—but disturbances from harvesting, fertilization, or irrigation cannot be fully accounted for.
• 3.Temperature-driven nonlinearity: Temperature fluctuations during sealed periods may cause slight deviations from linear concentration increases. A sampling window of 18:00–24:00 (or 20:00–21:00) is recommended to minimize this effect. Under unfavorable weather (e.g., rain or snow), keeping the greenhouse sealed and dark for ≥24 h allows verification of whether the selected window is representative. For manual bag sampling, at least four time points should be collected between 18:00 and 24:00, with linear regression achieving R² > 0.9; with more time points, the R² threshold can be relaxed.
• 4.Lack of spatial resolution: The method yields an integrated flux for the entire greenhouse. Compared with small soil chambers, it includes plant contributions and reduces the influence of fine-scale soil variability, but it cannot resolve within-greenhouse heterogeneity or compare emissions across subplots.
• 5.Applicability across systems: Whole-greenhouse flux estimation is accurate, labor-efficient, and cost-effective, but limited to sealed systems (plastic or glass). It cannot be applied in open-field settings, where micrometeorological methods are more appropriate. Automated small chambers are more versatile but are costly and highly sensitive to spatial variability.

Ethics statements

The authors declare that the present method complies with the journal’s requirements and does not involve human subjects, animal experiments, or data collected from social media platforms; therefore, ethics statements are not applicable.

CRediT author statement

Zhi Quan: Conceptualization, Methodology, Writing - original draft, Project administration, Funding acquisition. Xue Li: Investigation, Instrumentation, Data curation. Yi Zhu and Fulong Wei: Formal analysis, Validation, Field measurements. Yunting Fang: Project administration, Funding acquisition, Writing - review & editing.

The supplementary material includes time-series CO₂ and N₂O data from 15 Shouguang greenhouses measured with a Picarro G2508, with a 70-day continuous record from the first greenhouse.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Data availability

I have shared our data in the Supplementary material.