Plant materials

Ginseng seeds (which were morphologically and physiologically postmatured) were provided by the ginseng base in Tonghua County, Jilin Province, China. Seeds were sown in hole trays with one seed per hole in a greenhouse at Jilin Agricultural University (lat. 43°48′37.54″ N, long. 125°24′37.39″ E) on 26 May 2022; the trays comprised 21 holes of 6 × 3 × 10 cm (upper diameter × lower diameter × depth) and a seedling substrate of grass charcoal:vermiculite:perlite of 5:3:1 (volume ratio). The pH of the nursery substrate was 6.8 to 7.0, with a bulk density of 0.26 g·cm⁻³, 311.1 to 328.3 mg·kg⁻¹ alkaline-hydrolyzable nitrogen, 40.4 to 42.8 mg·kg⁻¹ available phosphorus, and 699.3 to 717.9 mg·kg⁻¹ available potassium. The light condition was natural light, the greenhouse air temperature was controlled at 15 to 25 °C, and the water content of the substrate was maintained at 50% to 60%. When the seedling leaves are completely spread flat, 36 trays of cavity tray seedlings with a uniform growth size were selected and randomly assigned to nine trays as a treatment group, with every three trays as a biological replicate; then, the selected ginseng cavity tray seedlings were sprayed with plant growth inhibitors.

Chemical treatment

We added CCC (Sinopharm Chemical Reagent Co., Ltd., Shanghai, China) to Tween as a surfactant and applied it by performing foliar spraying with a hand-held pressure sprayer at the leaf spreading stage of ginseng seedlings (23 Jun 2022) at a uniform time of 4:00 PM to mitigate the effect of high temperatures and strong light on the efficacy of the drug to the extent that the stems and leaves were completely wet and about to drip. The applied chemical concentrations were 100, 200, and 300 mg·L⁻¹, and the blank control group was sprayed with equal amounts of water.

Samples and data collection

The height and stem thickness of ginseng seedlings were measured on days 7, 14, 21, and 28 after foliar spraying of CCC. Nine plants were selected from each treatment group and there were three replications. The leaf area was measured with a foliar image analysis system (WinFOLIA; Regent Instruments, Quebec City, QC, Canada) that was accurate to 0.001 cm², and the fresh weight was measured by washing and drying the plants and weighing them with an electronic balance. Then, the plants were killed at 105 °C for 30 min and dried at 60 °C to a constant weight. The dry weight was measured with an accuracy to 0.001 g. The root-to-crown ratio was calculated.

Fresh ginseng seedling leaves were collected on days 7, 14, 21, and 28 after spraying, and the main leaf veins were removed and ground into powder with liquid nitrogen to determine the SOD, POD, CAT activity, and malondialdehyde (MDA) content. The SOD, POD, CAT, and MDA were determined by using corresponding kits (Suzhou Kemin Biotechnology Co.). The chlorophyll content and chlorophyll fluorescence parameters of ginseng seedling leaves were measured on day 28. The Chl a and Chl b contents were extracted by using an acetone–ethanol mixture (1:1), and the chlorophyll content was calculated according to the Lambert–Beer law. The absorbance values at 663, 645, and 470 nm were measured using an ultraviolet spectrophotometer (Model U-2900; Hitachi Co., Tokyo, Japan). The chlorophyll content (in mg·L⁻¹) was calculated as follows: chlorophyll a (Chl a) = 12.7D₆₆₃ − 2.69D₆₄₅; chlorophyll b (Chl b) = 22.9D₆₄₅ − 4.68D₆₆₃; and total chlorophyll t (Chl t) = 20.2D₆₄₅ + 8.02D₆₆₃.

The chlorophyll fluorescence parameters were measured by using an OS5-FL modulated chlorophyll fluorometer (Opti2science; Opti-science, Hudson, NH, USA) by selecting the middle leaflet of ginseng seedlings with three compound leaves. Light acclimation was measured as follows: real-time fluorescence F′ was measured after 15 min of endogenous action light [100 µmol/(m⁻²·s⁻¹)]; maximum fluorescence (Fm′) was measured after saturated pulsed light; and, finally, minimum fluorescence (Fo′) and photosynthetic electron transfer rate (ETR) were measured while the action light was off and the far-red light was on.

The stem ultrastructure observation was performed. Several pieces of 1-mm³ tissue were fixed in 2.5% glutaraldehyde for 24 h, poured into phosphate-buffered saline for 6 h, and fixed for 2 h after adding 1% osmium acid to the cut tissue. These pieces were dehydrated in 30% ethanol for 10 min, 50% ethanol for 10 min, and 70% ethanol uranium acetate (staining before embedding) for 3 h or overnight, 80% ethanol for 10 min, 95% ethanol for 15 min, 100% ethanol twice for 50 min, and epichlorohydrin for 30 min. After dehydration, the samples were first soaked in a solution prepared from propylene oxide and epoxy resin at a 1:1 volume ratio for 2 h, and then soaked in pure epoxy resin for 3 h. Next, they were embedded in pure epoxy resin in a 45 °C oven for 12 h and a 72 °C oven for 24 h. Thereafter, they were removed from the embedding block and trimmed after ultrathin sectioning (slice thickness, 70 nm; Leica UC-7 slicer; Leica Biosystems, Nussloch, Germany). Electron staining was conducted (lead staining) and a transmission electron microscope (JEM1400, Japan) was used to obtain photographs.

Data analysis

A completely randomized design was used in the experiment. All statistical analyses were performed using SPSS (version 25.0; IBM, Armonk, NY, USA) for the one-way analysis of variance and correlation analysis. Duncan’s test was used for the differential significance analysis (P < 0.05).

Effects of CCC on the growth index of ginseng seedlings

The CCC inhibited the growth of ginseng seedlings and significantly reduced the height of ginseng seedlings (Fig. 1A). The treatment groups with different concentrations on days 7, 14, 21, and 28 after CCC spraying significantly reduced the ginseng seedling plant height compared with that of the control group, which gradually decreased with the increased concentration applied and showed a negative correlation. The optimal concentration of CCC for high inhibition of ginseng seedlings was 300 mg·L⁻¹, which reduced the height of ginseng seedlings by 15.7% (day 7), 25.6% (day 14), 22.9% (day 21), and 24.0% (day 28) at different times after application, which proved that CCC has the most significant inhibitory effect on the height of ginseng seedlings on day 14 after application. On day 28 after spraying, the plant height of each treatment group decreased by 0.67 cm, 1.04 cm, and 1.44 cm, respectively, compared with that of the control group, and significant differences were apparent among all treatment groups (P < 0.05). Overall, ginseng seedlings showed the best inhibition of plant height growth after foliar spraying of CCC 300 mg·L⁻¹ at the leaf spreading stage. Plant growth regulators inhibited the increase of stem length but promoted the increase of stem thickness, and all CCC spraying treatments promoted an increase in the stem thickness of ginseng seedlings (Fig. 1B). On day 7 after application, each treatment group increased by 0.03 mm (100 mg·L⁻¹), 0.06 mm (200 mg·L⁻¹), and 0.04 mm (200 mg·L⁻¹), respectively, compared with the control group; however, this difference was not significantly different from that of the control group. On day 14, the stem thickness of the treatment group with CCC applied at 200 mg·L⁻¹ only was significantly higher than that of the control group, with an increase of 17.8% compared with that of the control group. By day 21 after application, the stem thickness of all treatment groups with CCC was significantly higher than that of the control group, which proved that CCC21 had the most significant effect on increasing the stem thickness of ginseng seedlings. On day 28, the efficacy of CCC weakened, and only the treatment group with the application of 200 mg·L⁻¹ had significantly higher stem thickness than that of the control group, with an increase of 7.2% compared with that of the control group. In general, the stem thickness of ginseng seedlings first increased and then decreased with the increasing CCC application concentration; it reached a maximum value at 200 mg·L⁻¹. The further growth of stem thickness was inhibited when the concentration continued to increase; therefore, CCC had the most significant effect on the growth of stem thickness of ginseng seedlings when foliar spraying of 200 mg·L⁻¹ was performed.

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The leaf length of ginseng seedling leaves of the concentration treatment group compared with that of the control group was significantly reduced (Fig. 2A). The leaf length decreased with increasing application concentrations and varied significantly between the treatment groups (Fig. 2A). A significant effect on leaf width was not observed when CCC was applied at 100 and 200 mg·L⁻¹, with decreases of 0.6% and 2.3%, respectively, compared with that of the control group. Significant differences were only observed when 300 mg·L⁻¹ was applied; the leaf width showed a significant decrease, proving that the concentration changes had less of an effect on leaf width. However, the effect on leaf length was observed much more clearly. With increasing application concentrations, significant decreases in the leaf area of 96.6% (100 mg·L⁻¹), 95.5% (200 mg·L⁻¹), and 86.1% (300 mg·L⁻¹), respectively, were observed in all concentration treatment groups compared with that of the control group, but there was no significant difference in the leaf area between the CCC 100 and 200 mg·L⁻¹ treatment groups (Fig. 2B). Plant growth regulators inhibit the growth of aboveground parts and promote the growth of belowground parts of ginseng seedlings. The fresh and dry weights of the aboveground parts decreased significantly with increasing application concentrations, and the most significant decrease was observed in the 300 mg·L⁻¹ treatment group, with 38.5% and 52.5% decreases in fresh and dry weights, respectively (Fig. 2C). The fresh weight and dry weight of underground parts first increased and then decreased with increasing application concentrations, and CCC reached the maximum value with the application of 200 mg·L⁻¹; the fresh weight was significantly higher than that of the control group, with an increase of 7.5% (Fig. 2D). The root length of ginseng seedlings decreased with the increasing application concentrations. Additionally, CCC at 100 mg·L⁻¹ significantly promoted the increase in the root length of ginseng seedlings compared with that of the control group, with an increase of 14.6%; there was no significant difference between the 200 and 100 mg·L⁻¹ treatment groups (Fig. 2E). The root-to-crown ratio of ginseng seedlings increased with increasing application concentration, and they were positively correlated, with the 200 and 300 mg·L⁻¹ treatment groups exhibiting significantly increased root-to-crown ratios compared with that of the control group by 29.9% and 58.8%, respectively (Fig. 2F). The CCC inhibited the increase in biomass of the belowground parts when applied at 300 mg·L⁻¹, whereas the root-to-crown ratio continued to increase, indicating that the higher concentration of the plant growth retardant affects the aboveground part of ginseng seedlings more than it does the underground part.

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Effect of CCC on the chlorophyll content of ginseng seedlings

Foliar application of CCC at 200 and 300 mg·L⁻¹ significantly promoted the Chl a content in the leaves of ginseng seedlings compared with that of the control group, with increases of 10.3% and 6.9%, respectively. The Chl b content in the leaves first increased and then decreased with increasing concentrations, with no significant difference between the 100 mg·L⁻¹ treatment group and the control group; however, that of the 200 and 300 mg·L⁻¹ treatment groups was significantly higher than that of the control group, with increases of 12.6% and 6.4%, respectively (Fig. 3A). After spraying CCC, the Chl t in each treatment group was 92.1% (100 mg·L⁻¹), 111.0% (200 mg·L⁻¹), and 106.8% (300 mg·L⁻¹) of the control group, with the 200 mg·L⁻¹ treatment group reaching the maximum (Fig. 3B). Overall, the foliar application of CCC at 200 mg·L⁻¹ to ginseng seedlings was able to significantly promote the accumulation of the chlorophyll content in the leaves of ginseng seedlings with increases of 10.3% (Chl a), 12.6% (Chl b), and 11.0% (Chl t), respectively, compared with that of the control group.

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Effect of CCC on chlorophyll fluorescence parameters of ginseng seedlings

Foliar spraying of ginseng seedlings with different concentrations of CCC promoted chlorophyll fluorescence parameters, maximum quantum efficiency of photosystem II (PSII) photochemistry (Fv/Fm), actual photochemical efficiency of PSII in the light [Y(II)], ETR, and photochemical quenching (qP) to varying degrees and decreased nonphotochemical quenching (NPQ) (Fig. 4). The Fv/Fm increased with increasing concentrations of CCC applications to 100.2%, 100.5%, and 101.5% of the control, and the 100 and 200 mg·L⁻¹ treatment groups were not significantly different from the control group; however, the 300 mg·L⁻¹ treatment group was significantly different from the control group (Fig. 4A). The trend of Y(II) was consistent with Fv/Fm, with 144.4%, 151.0%, and 154.4% increases compared to that of the control group for each treatment group, respectively; these values were significantly higher than that of the control group (Fig. 4B). The ETR first increased and then decreased with increasing CCC application concentrations and was significantly higher than that of the control group in all treatment groups, reaching a maximum in the 200 mg·L⁻¹ treatment group with a 152.8% increase compared to that of the control group (Fig. 4C). The NPQ values were significantly lower than that of the control group, at 85.8%, 68.6%, and 73.4% of the control group; the lowest NPQ value was reached in the CCC 200 mg·L⁻¹ treatment group (Fig. 4E). Overall, foliar spraying of CCC at 200 mg·L⁻¹ was most beneficial for improving the chlorophyll fluorescence parameters of ginseng seedling leaves and enhancing the photosynthetic capacity of the plants.

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Effect of CCC on the antioxi-dant system of ginseng seedlings

The CCC promoted the activities of the antioxidant enzymes SOD, CAT, and POD in ginseng seedlings to some extent, reduced the MDA content in the leaves, and alleviated the degree of membrane lipid peroxidation (Fig. 5). The SOD, CAT, and POD activities in ginseng seedlings showed a tendency to first increase and then decrease after CCC application, reaching the maximum value in each treatment group on day 14 after application; however, the MDA content showed the opposite trend, first decreasing and then increasing and reaching the minimum value on day 14 after application. The SOD activity of ginseng seedlings was significantly higher on day 7 after application, and all treatment groups were significantly higher than that of the control group. On days 14 and 21, the SOD activity of the 200 mg·L⁻¹ treatment group reached its maximum; these values were 126.4% and 113.4% that of the control group, respectively, and significantly higher than that the control group. The CAT activity of ginseng seedlings was improved by the CCC application, but only the 300 mg·L⁻¹ treatment group on day 14 and the 200 mg·L⁻¹ treatment group on day 28 were significantly different from the control group, with increases of 17.1% and 17.0%, respectively (Fig. 5A). The increase in POD activity of ginseng seedlings reached a significant effect on day 14, with increases of 40.0%, 47.9%, and 44.8%, respectively, compared with that of the control group, and the effect on the increase in POD activity had a long duration. By day 28, all treatment groups had significantly higher values than that of the control group, with no significant differences between treatment groups of different concentrations (Fig. 5C). The MDA content in the 100 mg·L⁻¹ treatment group was the lowest, with 80.6% and 73.5% that of the control group, respectively; these values were significantly lower than that of the control group. On days 21 and 28 after application, the 200 mg·L⁻¹ treatment yielded the best results, with values 26.4% and 20.7% lower than that of the control group, respectively (Fig. 5D). Taken together, the low and medium concentration applications were favorable for reducing the MDA content of ginseng seedlings plants, and the application of the high concentration (300 mg·L⁻¹) was most favorable for improving SOD, CAT, and POD activities; however, there were no significant differences compared with those of the low and medium concentration treatment groups. Therefore, the 200 mg·L⁻¹ application concentration was most favorable for improving the antioxidant enzyme activities and reducing cell membrane lipidation of ginseng seedlings.

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Effect of CCC on the microstructure of the stem of ginseng seedlings

Further study of the microstructure of the stem cells of ginseng seedlings showed that the number of intracellular starch grains was low (Fig. 6A), the number of stromal lamellae in chloroplasts was low and loosely arranged (Fig. 6B), and the cell wall thickness was small (Fig. 6C) in the control group; however, in the CCC-treated group, the number of intracellular starch grains increased significantly (Fig. 6D), the number of stromal lamellae in chloroplasts increased (Fig. 6D), the arrangement was more compact (Fig. 6E), and the cell wall thickness increased significantly (Fig. 6F).

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Effect of CCC on seedling growth and development

The effects of plant growth inhibitor application on the physiological traits of plants were exhibited by the shorter plant height, thicker stems, smaller leaves, shorter flower stems, and delayed flowering in aboveground parts as well as increased root length, more fibrous roots, and increased root weight in belowground parts (Chen et al. 2022; Kamran et al. 2018). On day 28 after application, although different concentrations of CCC treatment significantly reduced the height of ginseng seedlings by 11.2% to 24.1%, only the stem thickness of ginseng seedlings treated with 200 mg·L⁻¹ CCC significantly increased. Similarly, the effect of CCC applications on lettuce showed that foliar application of 500 mg·L⁻¹ did not change the plant stem strength or increase the diameter; only when the concentration reached 1500 mg·L⁻¹ could it significantly reduce the plant height and increase the stem diameter at the same time, which proved that CCC applications should reach a certain concentration to significantly affect the plant height as well as the stem diameter (Passam et al. 2008). The leaf size of ginseng seedlings decreased with increasing application concentrations, consistent with the conclusion that there is a linear relationship between increasing concentrations of plant growth retardant applications and reduced leaf areas of treated roses (Carvalho-Zanão et al. 2017), and that the reduction in the leaf area of ginseng leaves is mainly caused by significant inhibition of leaf length. Plant growth inhibitors can affect the redistribution and reuse of nutrients to varying degrees, promote a significant increase in the root length and weight of ginseng seedlings, inhibit the growth of aboveground biomass, and regulate the root-to-crown ratio of the plants, consistent with the results of studies of rapeseed blanket seedlings (Zuo et al. 2020) and passion fruit seedlings (Teixeira et al. 2021).

Although the application of plant growth inhibitors resulted in a smaller leaf area, it also resulted in a more intense green leaf color and increased chlorophyll content as well as improved the photosynthetic performance of the plant (Aphalo et al. 1997). Foliar application of CCC at a concentration of 200 mg·L⁻¹ was most effective for increasing the chlorophyll content of ginseng seedlings, with significant increases in Chl a and Chl b compared with that of the control group, significant increases in chlorophyll fluorescence parameters Y(II), ETR, and qP, and significant decreases in NPQ values. Previous research also showed that the chlorophyll content of CCC-treated lilies increased significantly (Zheng et al. 2012), thus further enhancing the photosynthetic capacity of the plants; the increase in the chlorophyll content was presumed to be attributable to the increase in the chlorophyll content as a result of the increased thickness of the plant leaves after CCC treatment (Tezuka et al. 1989). Another speculation was that CCC can increase chlorophyll content by inhibiting the synthesis of gibberellin acid. The CCC inhibits gibberellin biosynthesis at the stage of conversion of geranylgeranyl pyrophosphate to ent-kaurene; a plausible explanation for CCC blocking ent-kaurene is that it affects the enzymatic activity from geranylgeranyl pyrophosphate to ent-kaurene, with geranylgeranyl pyrophosphate conversion being blocked and converted more to diterpenes, which, in turn, are involved in carotenoid and chlorophyll synthesis (Rademacher 2000).

Chlorophyll fluorescence kinetics technology is widely used in photosynthesis mechanisms and plant stress resistance, and it is an ideal probe for studying the photosynthetic physiological status of plants and the relationship between plants and the stress of adversity (Durako 2012). Previous results have shown that some plant growth inhibitors can improve the chlorophyll fluorescence parameters of plants (Urfan et al. 2022). Both 2-diethylaminoethyl-3,4-dichlorophenylether (DCPTA) and CCC can improve chlorophyll fluorescence properties to varying degrees after being applied to maize, thus optimizing the upregulation mechanism of PSII reactions and enhancing photosynthetic capacity through increased chlorophyll content; furthermore, more chlorophyll molecules are used to increase the rate of electron uptake and transport (Wang et al. 2016). Similarly, our experimental results are consistent with those of previous studies that indicated that the foliar application of CCC to ginseng seedlings at a concentration of 200 mg·L⁻¹ resulted in an increased ETR by increasing the value of photochemical burst coefficients and reducing unnecessary nonphotochemical bursts. As a result, the goal of promoting an increase in the actual photochemical efficiency Y(II) of the plant PSII was finally reached.

The normal photosynthesis and respiration of plants are accompanied by the production of reactive oxygen species, and the accumulation of reactive oxygen species causes cell membrane lipid peroxidation and protein inactivation, which affect the normal growth and development of plants. The final decomposition product of membrane lipid peroxidation is MDA. The higher the activity levels of SOD, POD, and CAT, the better it is for maintaining the physiological balance in plants, resisting the adverse effects of external environmental factors, and enhancing the plant’s resilience (Pakzad et al. 2019). The results of previous studies showed that the plant growth inhibitor polyconazole was effective for enhancing the antioxidant enzyme activity of the plants after application to mango (Srivastav et al. 2010) and wheat (Nagar et al. 2021). The antioxidant enzyme activity was also significantly improved in CCC-treated maize. Our results have also shown that the SOD, POD, and CAT activities of ginseng seedlings are higher than that of the control after 28 d of 200 mg·L⁻¹ CCC treatment, and the POD and CAT activities increased to significant levels; additionally, the MDA content decreased significantly, and the plant resistance to stress was significantly improved.

Plant growth retardants inhibit the longitudinal growth of plant stems and promote lateral growth. The cell wall extensibility of plant stems was significantly and negatively correlated with the cell wall thickness. The thicker the cell wall was, the lower the plasticity and the less conducive it was to cell elongation (Nishijima 2023). Ophiopogon japonicus cell walls thickened after treatment with the plant growth retardant poloxamer, whereas cells decreased and were more tightly arranged (Zhang et al. 2021). Rice stem height decreased and strength increased after treatment with the plant growth retarder uniconazole because of an increased cell aspect ratio and smaller intercellular gaps (Lv et al. 2022). Our experimental results also showed an increase in stem thickness and a significant increase in stem strength of ginseng seedlings after CCC treatment. The stem changes were mainly caused by an increase in cell wall thickness and a decrease in the longitudinal length of the cells and their close alignment. Based on the submicrostructure, it was also hypothesized that the chloroplast stroma lamellae and starch grains were fewer in number in the control group because of the photosynthetic products mainly used to supply the cells with rapid elongation and growth. In contrast, after CCC treatment, the chloroplast stroma lamellae were stacked and tightly arranged, and the photosynthetic products tended to accumulate in the direction of assimilates because the longitudinal elongation of cells was inhibited because of the reduction in cell wall plasticity. As a result, the number of starch grains in chloroplasts was significantly higher than that of the control group.

Application and safety of CCC

One of the most extensively used plant growth regulators worldwide, particularly in cereal crops, is CCC (Wu et al. 2024). Despite the presence of residues in cereal crops, its usage has not been explicitly prohibited, largely because of its crucial role in mitigating lodging and ensuring optimal cereal crop yields. In regions such as the European Union, the United Kingdom, and Canada, CCC is sanctioned for application in cereal crops, notably wheat, oats, and barley. The European Food Safety Authority concluded that the proposed use of chlormequat on barley following good agricultural practices is unlikely to result in consumer exposure exceeding the toxicological reference values and, therefore, is unlikely to pose a risk to consumer health (Authority et al. 2020).

Initially, CCC was restricted to ornamental plant use in the United States. However, in Apr 2018, the US Environmental Protection Agency (EPA) published acceptable food tolerance levels of CCC in imported oat, wheat, barley, and some animal products, which permitted the inclusion of CCC in the US food supply. Subsequently, the permitted tolerance levels for oats were further augmented in 2020 (Temkin et al. 2024). Then, in 2023, the EPA proposed a decision for public comment and advocated for the inaugural inclusion of CCC in cereal crop cultivation, thereby equipping farmers with an additional means to enhance crop yields. Notably, the EPA’s human health risk assessments concluded that CCC poses negligible dietary, residential, and aggregate (combined dietary and residential exposure) risks to consumers (US Environmental Protection Agency 2023).

As a prominent agricultural producer, China permits the registration and application of CCC on maize, which ranks second in grain production, and wheat, which ranks third. Notably, there are comprehensive application examples and risk assessments for dual-use crops as well. Research conducted at the Hubei Academy of Agricultural Sciences revealed that the half-life of CCC in the stems of dual-use Artemisia selengensis Turcz. ranged from 6.9 to 9.1 d after application (Zheng et al. 2021). Moreover, the residue concentrations in stems after CCC spraying for 17 d at concentrations of 150 to 450 mg/kg remained below the limit standards stipulated in GB 2763-2021, thereby ensuring food safety (Zheng et al. 2021). Similarly, investigations conducted at the Ningbo Academy of Agricultural Sciences in Zhejiang Province demonstrated that treating Eleocharis dulcis, a dual-use medicinal and edible crop, with CCC not only augmented yield but also caused no significant alterations in taste (Ling et al. 2021). Moreover, chronic dietary risk assessments indicated that the CCC intake from Eleocharis dulcis remained within acceptable limits (Ling et al. 2021). Further bolstering the safety profile of CCC, research conducted at Changchun Normal University detected CCC residues in 10 of 12 samples from the primary ginseng production areas, with all of them below human health concern standards (Peng et al. 2021). Because ginseng is typically harvested after 4 to 6 years, and because the experiment used lower concentrations of CCC during the seedling stage, the safety and controllability of CCC application during ginseng cultivation were underscored.

Ensuring high-quality ginseng seedlings is paramount for achieving optimal yields and medicinal quality (Zhang et al. 2018). Although tray seedling cultivation can yield large-scale ginseng seedlings, issues like dense planting and inadequate air permeability may result in excessive elongation. Excessive elongation of the stem will cause the slender stem to fail to support the weight of the leaves. Consequently, the practicality of using CCC to control this elongation presents significant potential.

Because of ginseng’s status as a medicinal herb, we delved into assessing the impact of CCC on the active ingredient content of ginseng seedlings. Encouragingly, the findings revealed that CCC does not hinder the accumulation of active ingredient in ginseng seedlings within the same year. However, to comprehensively understand its effects, further investigations of how CCC influences the quality of ginseng medicinal materials at harvest age are warranted. Such research will provide valuable insights into optimizing the cultivation practices of this invaluable medicinal plant.

In summary, the results of this study showed that the CCC 200 mg·L⁻¹ treatment group had the best effect on significantly reducing the height of ginseng seedlings, the most significant effect on increasing the stem thickness, root length, and root weight, and the least inhibitory effect on the increase in the leaf area of ginseng leaves. The height of ginseng seedlings decreased by 17.4%, stem thickness increased by 7.4%, leaf area decreased by 4.5%, root length increased by 6.2%, and root-to-crown ratio increased by 29.8%. The POD activity increased by 38.7%, the CAT activity increased by 17.0%, and the MDA content decreased by 18.1%. Stem strength was enhanced, and the plant’s resistance to collapse improved by adjusting the cell gap and increasing the cell wall thickness. Therefore, it is recommended to control the growth of ginseng seedlings, adjust the plant stature, and improve the plant resistance to stem collapse and stress by foliar spraying CCC at 200 mg·L⁻¹ during production.