**1. Introduction**

Rivers connect terrestrial and marine ecosystems and are important channels for the transfer and transformation of nutrient elements. The transport of particulate and dissolved matter from rivers to the ocean is of great significance for the material cycle of the ecosystem, and it is estimated that the total amount of particulate sediment and dissolved matter transported into the ocean by rivers worldwide is 15.5 × <sup>10</sup><sup>9</sup> t/a and <sup>4</sup> × 109 t/a, respectively [1]. At the same time, to make full use of water energy resources, water conservation has been pursued around the world in recent decades. The construction of dams transforms a single river ecosystem into a river–reservoir ecosystem, river continuity is forced to change, and the pattern of material transport from the source to the estuary also changes. Reservoirs are usually characterized by poor flow, water temperature stratification, and large depth, and the retention and transformation of various substances in reservoirs are relatively considerable. Many studies have also been carried out on the biogeochemical cycle of river–reservoir ecosystems [2–5].

As essential elements for the growth of life, carbon, and nitrogen play an important role in ecosystems. Related studies have shown that carbon and nitrogen can reflect the construction of aquatic food chains and the division of trophic levels in aquatic ecosystems [6] and can also be used for carbon and nitrogen isotope analysis in aquatic plants and plankton [7,8]. Water bodies in karst areas are generally rich in calcium and somewhat

**Citation:** Zhou, Z.; Kong, J.; Zhang, F.; Zou, Y.; Xie, J.; Wen, C. Study on the Carbon and Nitrogen Isotope Characteristics and Sources and Their Influence on Carbon Sinks in Karst Reservoirs. *Land* **2023**, *12*, 429. https://doi.org/10.3390/ land12020429

Academic Editor: Tilo Ziehn

Received: 1 January 2023 Revised: 31 January 2023 Accepted: 3 February 2023 Published: 6 February 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

alkaline. In terms of the impact of nutrients on water quality, even if the concentrations of nitrogen and phosphorus are low, aquatic plants (blue-green algae) and plankton can develop through the carbon fertilization effect. The input of exogenous nitrogen will contribute to the growth of living organisms such as microorganisms and plants in water, which will affect the absorption and decomposition of carbon [3,4].

However, increasing human activities such as land-use change, fossil fuel burning, and agricultural production have altered the natural state of the carbon and nitrogen cycle [9,10]. For example, the use of nitrogen fertilizer in agriculture can promote photosynthesis by increasing the net primary productivity of vegetation and microbial activities and increase the decomposition of organic matter by microorganisms, but this will produce more CO2, part of which is re-discharged into the atmosphere as a carbon source, and the other part infiltrates into the karst surface zone and vadose zone with rainfall to form carbonic acid, increasing the dissolution of carbonate rocks [11,12]. Relevant studies also show that nitric acid produced by agricultural and urban activities interferes with the karst carbon cycle [13–15]. In contrast to carbonate rocks dissolved by carbonic acid, carbonate rocks dissolved by nitric acid do not consume CO2 in the atmosphere/soil, leading to an increase in HCO3 <sup>−</sup> and (Ca2+ + Mg2+) concentrations in water, playing the opposite role in CO2 emission reduction [16,17]. Baker et al. (2008) showed that a river flowing through the city had the highest dissolved inorganic carbon (DIC) concentration in the carbonate rock area of Britain [18]. Barnes et al. (2009) found that DIC was higher in watersheds dominated by urban land than in watersheds dominated by forestland [13]. It is estimated through laboratory simulation that fertilization of cultivated land in karst regions will lead to an additional increase in the (Ca2+ + Mg2+) concentration in rivers by 5.7 × <sup>10</sup><sup>12</sup> mmol/a and will release CO2 to the atmosphere [19]. A study in a karst basin in southwest France found that the application of chemical fertilizers increased the concentration of nitric acid in river water and estimated that the amount of atmospheric CO2 absorbed by the weathering of carbonate rocks in the basin decreased by 7–11%, and the karst carbon sink decreased by 5.7–13.4% in the whole region of France [20]. According to Brunet et al. (2011), the nitric acid formed by nitrification of nitrogen fertilizer can cause soil and water acidification, increase the concentration of alkaline cations, change the carbon budget, and actively participate in the weathering of carbonate rocks [21]. A study in the typical karst agricultural area of Southwest China shows that H+ released from the nitrification of nitrogen fertilizer accelerates the weathering of carbonate rocks, which not only reduces the consumption of atmospheric CO2 but also increases the HCO3 − flux by approximately 20% [22].

Southwest China has the largest contiguous distribution of carbonate rocks in the world, with an exposed area of carbonate rocks of 54 × <sup>10</sup><sup>4</sup> km2 [23]. Southwest China has also become a key area for studying carbon and nitrogen cycling. The Pingzhai Reservoir is located in Southwest China, and its water transfer scope involves the Yangtze River basin and the Pearl River basin [24]. Its carbon and nitrogen concentrations are of great significance for the water quality security of the Yangtze and Pearl Rivers. In terms of geological background, the Pingzhai Reservoir is located in a deep river canyon, with very thick carbonate rock strata distributed on both sides of the canyon. Affected by karstification, a multilayered karst hydrogeological structure has been formed. The karst morphology mainly includes peak-cluster depressions, dissolving gullies and troughs, falling caves, funnels, karst caves, and karst pipeline systems. In the region, in recent years, with the increase in population, frequent industrial and agricultural activities in the basin, and the large amount of agricultural fertilizers used with low utilization efficiency [24,25], nutrients have entered rivers and reservoirs through cracks in the karst and underground rivers along with the runoff and pore water generated by precipitation, leading to the accumulation of nitrogen nutrients and participating in water-rock interactions. Previous studies on carbon and nitrogen in the Pingzhai Reservoir and its inflow river were relatively isolated [25–27], but with the progress of research, it has been found that it is very important to explore the coupling of carbon and nitrogen and its environmental effects. The objectives of this study were to explore the water hydrochemical types and the spatial and temporal

distribution characteristics of nitrate nitrogen and oxygen isotopes and dissolved inorganic carbon isotopes in a karst reservoir basin, determine the sources of DIC and nitrate in water, explore the carbon and nitrogen coupling cycle in different periods, and quantitatively evaluate the impact of the carbon and nitrogen coupling cycle and nitric acid from external sources on carbonate dissolution in the basin. This study can provide a reference for the study of the effect of C–N coupling and karst carbon sinks on the river–reservoir continuum in karst regions.

#### **2. Materials and Methods**

## *2.1. Overview of the Study Area*

The Pingzhai Reservoir (105◦17 3 E–105◦26 44 E, 26◦29 33 N–26◦35 38 N) is the source reservoir of Guizhou's Central Water Control Project and undertakes the functions of irrigation, drinking water supply, and power generation in the region (Figure 1). The reservoir is formed by the convergence of five rivers (Nayong River, Shuigong River, Zhangwei River, Baishui River, and Hujia River) in the upper reaches of the reservoir, and the drainage area is 833.77 km2. The construction of the reservoir was completed in 2015, the maximum dam height is 157.5 m, the maximum water level is 1331 m, the regulated storage capacity is 448 million m3, and the total storage capacity is 1.089 billion m3. The study area is located in the subtropical monsoon climate zone, summer is hot and rainy, winter is mild and slightly rainy, the annual average temperature is 14 ◦C, the annual average rainfall is between 1200 and 1500 mm, and the rainfall has seasonal differences under the influence of the monsoon climate. The wet season is from May to August, the dry season is from November to February, and the normal season is March, April, September, and October.

**Figure 1.** Overview map of the study area ((**a**) is the chronostratigraphic diagram and sampling points distribution of the study area; (**b**) is the DEM of the study area; (**c**) is the reservoir and dam; (**d**) is the land use types map).

The terrain and geomorphology of the study area are complex and belong to the middle-low mountain valley landform of tectonic dissolution and erosion. The geological structure pattern is controlled by the Yanshan movement, and anticlines, synclines, and compressive faults are formed. As shown in Figure 1, the main outcrops are the Permian Dalong Formation (P*3d*), Longtan Formation (P3*l*), Maokou Formation (P2*m*), Triassic Yongningzhen Formation (T1*yn*), Yelang Formation (T1*y*), and Guanling Formation (T2*g*). The stratigraphic lithology is carbonate rocks such as limestone and dolomite and clastic rocks such as sandy mudstone, shale intercalated marl, and coal. The main land-use types in the study area were cultivated land and forestland, followed by grassland, construction land, and unused land. Affected by regional lithology and crop cultivation habits, the main soil types are yellow-brown soil, lime soil, and yellow soil, in addition to a small amount of paddy soil. The main crops are rice, corn, and potato. The main fertilizers used in the region are ammonium-based N fertilizer (urea), nitrogen–phosphorus compound fertilizer, and animal manure; the applied pesticides mainly include insecticides and rust removers, which pose the risk of agricultural nonpoint source pollution.
