Residual effect interval nicosulfuron applications and soybean sowing: Initial tolerance and agronomic impacts

Abstract

Context:  The persistence of herbicides in the soil hinders management of subsequent crops, affecting emergence and initial development. Nicosulfuron stands out among such herbicides, with potential for use in pre-sowing soybeans.

Objective:  To evaluate the initial tolerance of soybean varieties to nicosulfuron applied at different intervals before sowing.

Methods:  Two experiments evaluated the tolerance of soybean varieties to nicosulfuron. The first was conducted in a greenhouse in a completely randomized design with five replications (3x6 factorial). Factor 1 included two application times: 28 and 7 days before sowing (DBS) and a control, while factor 2 comprised six varieties: Olimpo IPRO, Iguaçu IPRO, Guepardo IPRO, M7601 i2x, Tormenta CE and ST700 i2x. In turn, phytotoxicity, biometric characteristics, fluorescence and dry mass were evaluated. The second experiment was conducted in the field in a randomized block design with six replications (2x4+1 factorial). Factor 1 included the Olimpo IPRO and Iguaçu IPRO varieties, and factor 2 the application times (7, 14, 21 and 28 DBS), in addition to the control. Plant stand, height, stem diameter, chlorophyll a and b index, and yield were evaluated.

Results:  Nicosulfuron application before sowing affected the initial development of the varieties differently. The Guepardo IPRO, ST700 i2X, and Tormenta CE varieties showed greater tolerance. There was no significant effect of the interaction between variety and application time in the field.

Conclusion:  Soybean varieties show differences in tolerance to nicosulfuron; choosing more tolerant genotypes reduces the risk of observing deleterious effects on the crop.

Keywords:
Selectivity; Herbicide Residues; Varietal Sensitivity; Pre-sowing; Early Development; Glycine max

1. Introduction

The agronomic performance of the soybean crop, as well as yield and grain quality, can be compromised by several factors, among which the interference imposed by weeds stands out (Forte et al., 2017). Weeds benefit from the interference process because they exhibit high hardiness, resistance to pests and diseases, the ability to produce a large number of viable seeds, ease of seed dispersal, and rapid transition from the vegetative to the reproductive phase, among several other factors inherent to these species (Silva et al., 2016). The aggressive characteristics provide weeds with advantages in the competition process with agricultural crops, resulting in yield losses and consequent damage to the farmer (Silva et al., 2016).

The degree of weed interference in competition with the crop depends on factors related to the weed community (floristic composition, density, frequency and dominance), crop (variety, spacing and seeding density), environment (climate, soil and management) and the coexistence period between weeds and the crop (time and duration) (Castro et al., 2019), where studies in the literature indicates that weeds can cause soybean grain production losses of close to 90% if left uncontrolled. Integrated weed management (IWM) is considered the main tool for reducing problems related to the weed community interfering in the crop of interest. IWM is based on integrating control methods, making cropping systems unfavorable to weeds, minimizing the ability of these species to survive the imposed control practices (Nunes et al., 2010). Chemical control is the most widely adopted among the different methods which compose IWM because it is economically accessible, enables a reduction in labor, and allows for rapid application when compared to the mechanical method (e.g., weeding).

With the development of glyphosate-tolerant materials, an herbicide called Roundup Ready® (RR®) has become the most widely used in grain crops. RR® technology has had a significant impact on Brazilian agriculture, especially for weed management in soybean crops. It initially favoured management of difficult-to-control species, reducing production costs, yield losses, and simplifying operations in the fields (Silva et al., 2021). However, the excessive use of glyphosate in production systems favoured selection of biotypes resistant to this herbicide (Silva et al., 2021). Due to the problem of glyphosate-resistant weeds, it is essential to rotate the herbicides used for chemical control of the weed community.

In this context, herbicides from the sulfonylurea chemical group are important and should be considered for weed control and preventing selection of glyphosate-resistant biotypes (Silva et al., 2018). Sulfonylurea herbicides act on the synthesis of essential amino acids for plants, blocking the synthesis of valine, leucine, and isoleucine by inhibiting the enzyme acetolactate synthase (ALS) or acetohydroxyacidosynthase (AHAS), which catalyzes two reactions in parallel (Rodrigues; Almeida, 2018).

Nicosulfuron stands out among these herbicides in the sulfonylurea chemical group. It is characterized as a post-emergence, systemic, and selective active ingredient for maize crops, presenting a broad action spectrum, although it is more effective in controlling grasses (Poaceae) (Rodrigues; Almeida, 2018). With a pKa of 4.3, nicosulfuron behaves as a weak acid in most soils. Its solubility and hydrophilic character are influenced by the pH of the medium: the Kow value at pH 5 is 0.44; the value at pH 7 is 0.018; and the Kow value at pH 9 is 0.0068. Its mobility in the soil is very low, with an average Koc of 30 mL g^-1 of soil at pH 6.5. Its persistence in the soil varies according to pH, but the average half-life of this herbicide in the field is 21 days at pH 6.5 (Rodrigues; Almeida, 2018).

The persistence of herbicides in the soil and their carryover potential are mainly influenced by interactions between herbicide molecules and components of the soil environment, such as microbiotas, microfauna, organic matter, and soil temperature, as well as factors such as pH, moisture, and oxygen availability (Mendes et al., 2017). Residual herbicides are characterized by having a longer period of activity in the soil. However, these compounds can result in a residual effect (carryover), which can cause negative environmental impacts. The carryover potential depends on the herbicide used, the crop in succession, and the environmental conditions after herbicide application (Mancuso et al., 2011).

Sulfonylurea-tolerant soybean (STS®) was developed using the seed mutagenesis technique with the alkylating agent ethyl methanesulfonate (EMS) and is not a trait (transgenic) (Silva et al., 2016). The EMS agent does not cause mutation by insertion into DNA, but rather by modifying an existing base, introducing an alkyl radical (in this case, ethyl) frequently found in the nitrogenous base guanine (Rogozin et al., 2001; Greene et al., 2001). The Als1 allele provides soybean varieties with tolerance to chlorimuron, nicosulfuron, rimsulfuron, sulfometuron, tifensulfuron, tribonuron, and flucarbazone herbicides, while the Als2 allele promotes tolerance to these herbicides and to imazapyr herbicide. Therefore, in the present study, soybean cultivars with and without STS® technology were used to compare the differential response to the application of the herbicide nicosulfuron. It is assumed that soybean cultivars with STS® technology exhibit greater herbicide tolerance compared to non-STS® cultivars, resulting in lower levels of phytotoxicity and less impact on agronomic performance.

In this context, the objective of this study was to evaluate the selectivity of nicosulfuron herbicide applied in periods prior to sowing soybean varieties in both the initial tolerance of the crop and in its agronomical performance.

2. Material and Methods

2.1 Greenhouse assay

The experiment was conducted from May to July 2024 in a climate-controlled greenhouse (Van der Hoeven, Double Poly Pad Fan model), with temperatures ranging from 18 to 29°C, in the weed laboratory located on the premises of the Federal Institute of Education, Science and Technology Goiano, Rio Verde Campus (17° 48' 18" S, 50° 54' 15" W and altitude of 744 m), in Rio Verde (Goiás), Brazil.

The substrate used to fill the pots was prepared from soil classified as Latossolo Vermelho, obtained from areas with no history of herbicide application in the municipality of Rio Verde. The physicochemical composition was pH (CaCl2) Un. 5.0, Ca mmolc dm^-3 22.5, Mg mmolc dm^-3 11, Ca+Mg mmolc dm^-3 33.5, Al mmolc dm^-3 3.5, H+Al mmolc dm^-3 50, CEC mmolc dm^-3 86.35, P (Resin) mg dm^-3 8, K mmolc dm^-3 2.85, S-SO4 mg dm^-3 10.5, B mg dm^-3 0.15, Cu mg dm^-3 2.9, Fe mg dm^-3 32.5, Mn mg dm^-3 12.75, Zn mg dm^-3 0.6, organic matter 32.5%, Al Sat. (M%) 9%, Base Sat. (V%) 36.35%, clay 44%, silt 12% and sand 44%. In turn, 5-liter pots were filled with the substrate to approximately 0.5 cm from the rim, with each pot constituting an experimental unit.

The adopted experimental design was completely randomized with 5 repetitions, with the treatments arranged in a 3 x 6 factorial arrangement. The first factor consisted of two application times of nicosulfuron before soybean sowing: 28 and 7 days before sowing (DBS), plus a control without herbicide application. A dose of 60 g a.i. ha^-1 of nicosulfuron herbicide was applied (Accent^®, Corteva Agriscience). The second factor consisted of six soybean varieties, namely: Olimpo IPRO, Iguaçu IPRO, Guepardo IPRO, M7601 i2x, Tormenta CE and ST700 i2x. The characteristics of the cultivars used, including cycle, maturity group, and growth habit, are presented in Table 1.

Table 1
Growth habits, maturity group, technology, company holding the varieties used in this study

Varieties	Growth habit	Maturation group	Technology	Holding company	Tolerance to sulfonylureas (STS)
Guepardo IPRO	Indetermined	6.7	Intacta RR2 PRO®	BRASMAX	Yes
Olimpo IPRO	Indetermined	7.7	Intacta RR2 PRO®	BRASMAX	No
Tormenta CE	Indetermined	7.6	Conkest Enlist®	BRASMAX	Yes
Iguaçu IPRO	Indetermined	6.4	Intacta RR2 PRO®	SEEDCORP HO	Yes
M7601 i2x	Indetermined	7.6	Intacta 2 Xtend®	MONSOY	No
ST700 i2x	Indetermined	7.0	Intacta 2 Xtend®	SOYTECH	No

The application was performed on 07/05/2024 to ensure same-day sowing representing a 28 DBS interval, and the other treatments were applied on 28/05/2024 after simulating a 7 DBS interval. The applications were done with a CO2-pressurized backpack sprayer, with a constant pressure of 2 bar and an application rate of 150 L ha^-1, equipped with a 4-nozzle TT110.02 boom, spaced 0.5 m apart. Meteorological conditions were monitored at the application time, with average temperatures of 20.8°/27.8°C, relative humidity (RH) of 74.6/61.2%, and wind speed of 4.1/6.6 km h^-1. Soybean sowing was done manually on 06/04/2024, with 4 seeds sown per experimental unit. Irrigation was done manually twice a day to keep the soil moist.

Plant height was measured at 14 DAE using a tape measure. Stem diameter was assessed at 21 DAE using a digital caliper, with 4 plants being measured horizontally. The fluorescence reading was performed on the same day, when the plants were in the V2/V3 phenological phase. In the evaluation, the most expanded leaf was adapted to the dark with the aid of tweezers for approximately 30 minutes. After this period, the data were read using the Fluorpen FP100 instrument. In this evaluation, the following were determined: quantum efficiency of Photosystem II (Fv/Fm), quantum yield of electron transport (Phi_Eo), quantum yield of dissipation energy (Phi_Do), absorption-based performance index (Pi_Abs), energy flux absorbed per active reaction center (ABS/ RC), energy flux trapped by active PSII (TRo/RC), electron transport flux per reaction center (ETo/RC), and energy flux dissipated (as heat and fluorescence) by active PSII (DIo/RC). In addition, the plants were thinned, with two plants remaining per pot.

The final plant height and root size were evaluated at 28 DAE using a tape measure, a shovel, and pruning shears to carefully remove the roots. A cut was made at the base of the soil, and the shoot height was measured. Furthermore, roots were removed from the soil with a shovel and their size was assessed.

Finally, the shoot was placed in one bag and the root part in another in order to measure the dry mass of both parts. These bags were placed in a forced-air oven at 65°C for 72 hours, after which they were weighed on an analytical scale.

2.2 Field assay

The experiment was conducted under field conditions during the 2023/2024 growing season, from November 2, 2023 to February 28, 2024, at the COMIGO Technological Center, located in the municipality of Rio Verde, Goiás (17°45'42" S, 51°02'09" W and an altitude of 827 m). The region's climate is classified as Aw, tropical savanna, according to the Köppen climate classification, with rain in the summer and drought in the winter. The accumulated rainfall during the experimental period is shown in Figure 1, according to data from the automatic weather station located at the COMIGO Technological Center in Rio Verde, Goiás (S 17°45'45" W 51°02'16").

Figure 1
Data on accumulated precipitation, temperature, and average relative humidity during the experiment period.

The soil in the experimental area is classified as a Latossolo Vermelho distrófico. An analysis of soil samples collected at a depth of 0 to 20 cm was performed before the experiments were set up, which revealed the following physical-chemical properties: pH in CaCl2 of 6.0; 2.31 cmolc dm^-3 of H⁺+Al⁺³; 4.84 cmolc dm^-3 of Ca⁺²; 1.13 cmolc dm^-3 of Mg⁺²; 63.4 mg dm^-3 of K; 70.19 mg dm^-3 of P; 19.68 g dm^-3 of O.M.; 47.61% sand; 16.62% silt; 35.77% clay (clayey-sandy texture); and V% (Base saturation): 72.6; CEC: 8.44.

The pre-plant burndown of the area was performed chemically before the experiment was set up with glyphosate herbicide (Zapp QI®, Syngenta) at a dose of 1250 g a.e ha^-1 to eliminate weeds. Mechanized sowing was subsequently performed using a no-till system on November 2nd, 2023. Approximately 18 seeds per meter were planted for the Iguaçu IPRO variety, and 13 seeds per meter for the Olimpo IPRO variety. Cultural practices, such as pest and disease management, were conducted according to the needs and technical recommendations for the crop.

The experimental design used was a randomized block design with six replications under a factorial scheme (2 x 4) + 1. The first factor consisted of the Olimpo IPRO and Iguaçu IPRO soybean varieties, while the second factor consisted of the different application times and sowing interval of the soybean, with these evaluations being conducted at 7, 14, 21 and 28 DBS. The additional treatment consisted of the control, which did not receive the nicosulfuron application. A dose of 60 g a.i. ha^-1 was used in the nicosulfuron herbicide application (Accent^®, Corteva Agriscience).

The herbicide applications were performed using a CO₂-pressurized research sprayer with an INPI patent (BR 102016007565-3), mounted on a tractor (LS 60, 60 hp, Landini). The spray bar was equipped with ten nozzles spaced 0.5 m apart, maintained at a height of 0.5 m from the vegetation cover. The working pressure on the air-induction double-flat jet spray nozzles (ADIA/D 110015; Magnojet) was 300 kPa (43.5 psi) and the application volume was 150 L ha^-1. The applications were carried out at seven-day intervals. The first application occurred on October 5, 2023, between 4:10 PM and 4:32 PM, under environmental conditions characterized by an air temperature of 31.7°C, RH of 48.35%, and wind speed of 1.0 km h^-1. The second application was carried out on October 12, 2023, from 9:12 AM to 9:36 AM, with a temperature of 27.9°C, RH of 60.85%, and wind speed of 1.25 km h^-1. The third application occurred on October 19, 2023, between 8:21 AM and 8:45 AM, with a recorded temperature of 31.5°C, RH of 53.40%, and wind speed of 0.85 km h^-1. The fourth application was carried out on 10/26/2023, from 10:28 to 10:54, under a temperature of 26.7°C, RH of 74.50% and wind speed of 0.6 km h^-1.

The evaluations began 14 DAE of the plants. At that time, the number of plants was counted to determine the initial stand. A tape measure was used for this evaluation, with which 4 m² was measured in two central lines. Then, the number of plants present in this area was counted. The height of five central plants of each variety was also subsequently measured using a tape measure, based on the soil surface and the last fully expanded leaf of the plant. Stem diameter was assessed at 21 DAE using a digital caliper 5 cm from the ground, on five random plants of each variety.

Next, the relative chlorophyll a and b index was evaluated at flowering using a Falker CFL1030 chlorophyll meter.

The device was calibrated before starting the evaluation to obtain more accurate results. In turn, five plants were randomly selected for this analysis, and the measurement was taken on the central leaflet of the third fully expanded trifoliate leaflet from the apex to the base of the plants.

Yield was evaluated at the end of the crop cycle. To do so, three central rows of 3 meters were harvested. After harvesting these rows (totalling 9 m²), the soybeans were threshed, and then the grains were cleaned and weighed, and the moisture content was corrected to 13%. The final plant stand was also evaluated by counting the number of plants in two central rows of 4 m². In addition, five plants were randomly harvested from the plot to assess several indices. In turn, plant height, number of pods per plant, number of grains per pod, and thousand-grain weight were evaluated using a measuring tape when necessary.

2.3 Statistical analysis

Prior to performing the analysis of variance, the data were subjected to verification of the basic assumptions (normality of errors and homogeneity of variances). Both basic assumptions were met for the different response variables analyzed, eliminating the need for data transformations. The statistical analysis used in this study was performed using the SISVAR statistical software. The data were subjected to analysis of variance using the F-test (p≤0.05), and the means were compared using Tukey's test for the greenhouse experiment and the Student's t-test (LSD) for the field experiment when a significant effect was found between the interaction of the evaluated factors or within an isolated factor, adopting a 5% probability level for both.

3. Results and Discussion

3.1 Greenhouse assay

The results of the greenhouse assay (Table 2) demonstrate that the nicosulfuron herbicide application prior to sowing significantly impacted the initial development of soybean varieties, with effects varying according to the application time and genetic material.

Table 2
Plant height, stem diameter, root length, shoot dry mass (SDM) and root dry mass (RDM) of soybean varieties subjected to nicosulfuron application at two times prior to sowing

Varieties	Height at 14 DAE (cm)			Height at 28 DAE (cm)
	28 DBS	7 DBS	Control	28 DBS	7 DBS	Control
Guepardo IPRO	10.06 Aa	9.40 Aa	10.03 Aab	15.23 Aa	13.57 Aa	13.53 Aab
Olimpo IPRO	8.79 Bab	9.75 Aba	10.45 Aab	12.17 Bbc	12.72 ABabc	14.80 Aa
Tormenta CE	7.21 Bb	7.22 Bb	9.06 Ab	9.62 Ac	10.35 Ac	11.18 Ab
Iguaçu IPRO	10.50 Aa	10.11 Aa	11.72 Aa	12.85 Aab	13.46 Aab	15.15 Aa
M7601 i2x	8.94 Aab	6.99 Bb	10.18 Aab	10.25 Bbc	9.83 Bc	13.71 Aab
ST700 i2x	10.06 Aa	9.93 Aa	11.12 Aa	12.47 Babc	10.60 Bbc	15.98 Aa
CV (%)	10.12			11.09
LSD Column	2.01			2.92
LSD Line	1.64			2.38

	Diameter at 21 DAE (mm)			Root length at 28 DAE (cm)
	28 DBS	7 DBS	Control	28 DBS	7 DBS	Control
Guepardo IPRO	2.30 Aa	2.49 Aab	2.40 Aab	18.70 Aab	22.81 Aa	18.62 Aab
Olimpo IPRO	2.58 Aa	2.83 Aa	2.62 Aa	18.18 Aab	21.87 Aa	19.37 Aab
Tormenta CE	2.49 Aa	2.51 Aab	2.10 Bb	24.31 Aa	20.25 Aa	26.25 Aab
Iguaçu IPRO	2.38 Aa	2.56 Aab	2.45 Aab	12.68 Bb	12.12 Ba	27.93 Aa
M7601 i2x	2.39 Aa	2.43 Ab	2.49 Aa	17.57 Aab	14.15 Aa	17.06 Ab
ST700 i2x	2.52 Aa	2.51 Aab	2.49 Aa	20.50 Aab	13.86 Aa	20.68 Aab
CV (%)	6.84			26.69
LSD Column	0.35			10.75
LSD Line	0.28			8.77

	SDM at 28 DAE (g)			RDM at 28 DAE (g)
	28 DBS	7 DBS	Control	28 DBS	7 DBS	Control
Guepardo IPRO	0.79 Aa	0.65 Aa	0.50 Ab	0.44 Aa	0.30 ABb	0.19 Bb
Olimpo IPRO	0.47 Ba	0.43 Bab	0.79 Aab	0.23 Bb	0.24 Bb	0.56 Aa
Tormenta CE	0.52 ABa	0.45 Bab	0.77 Aab	0.23 Ab	0.26 Ab	0.25 Ab
Iguaçu IPRO	0.54 Aa	0.22 Bb	0.76 Aab	0.27 Bab	0.54 Aa	0.50 Aa
M7601 i2x	0.49 Aa	0.35 Aab	0.51 Ab	0.19 Ab	0.16 Ab	0.29 Ab
ST700 i2x	0.61 Ba	0.56 Bab	0.96 Aa	0.45 Aa	0.27 Bb	0.39 ABab
CV (%)	30.22			30.50
LSD Column	0.37			0.20
LSD Line	0.29			0.16

• * Means followed by different lowercase letters in the Column and uppercase letters in the Row differ from each other by Tukey's test (p≤0.05). DAE: days after emergence; DBS: days before sowing.

The Tormenta CE and M7601 i2x varieties showed a significant reduction regarding plant height evaluated at 14 DAE when the herbicide was applied at 7 DBS. In addition, the Olimpo IPRO and Tormenta CE varieties also exhibited shorter plants compared to their controls in the application at 28 DBS. The effects persisted for the Olimpo IPRO (application at 28 DBS), M7601 i2x, and ST700 i2x varieties (both times) at 28 DAE, which showed reduced height. The Tormenta CE variety was initially sensitive, but recovered and equaled the control in this subsequent evaluation. The comparison between varieties showed that Tormenta CE and M7601 i2x consistently presented the lowest heights, both under the effect of the herbicide and in the control, indicating a genotypic influence on plant architecture, as reported by Kim et al. (2022). The initial reduction in height can be attributed to the nicosulfuron action, an inhibitor of the ALS enzyme, which is essential for the biosynthesis of branched-chain amino acids, compromising cell division and expansion (Peng et al., 2024). The persistence of the effect in some varieties (M7601 i2x and ST700 i2x) and the recovery of others (Tormenta CE) suggest differences in metabolic sensitivity or detoxification capacity among genotypes, possibly related to the activity of enzymes such as cytochrome P450 and glutathione S-transferase (Wang et al., 2022; Bakaeva et al., 2024; Liu et al., 2024).

In turn, most varieties were not significantly affected by the herbicide regarding their stem diameter at 21 DAE. The exception was the Tormenta CE variety, which showed a smaller diameter in the control compared to the herbicide treatments. This suggests that nicosulfuron did not compromise stem thickness growth for most genotypes, with this parameter being associated with structural support and water and nutrient conduction, which is less sensitive to chemical stresses in tolerant varieties (Swaef et al., 2015).

Root length was only significantly affected in the Iguaçu IPRO variety, which had its roots shortened by herbicide application at both times, reducing from 27.93 cm (control) to approximately 12 cm. This result indicates a specific impact of herbicide residue on the root system of this variety, possibly due to ALS inhibition in the meristematic regions of the root, which are essential for cell elongation (Kaur et al., 2024). This effect may compromise water and nutrient absorption, affecting agronomic performance (Guan et al., 2020).

The shoot dry mass (SDM) was reduced by the herbicide in several varieties. Olimpo IPRO and ST700 i2x stood out, showing lower SDM in applications compared to the control. The Iguaçu IPRO variety showed a significant reduction when applied at 7 DBS. Root dry mass (RDM) was also affected, with Guepardo IPRO and ST700 i2x varieties exhibiting higher RDM in the 28 DBS application, while Iguaçu IPRO stood out in the 7 DBS application. Olimpo IPRO and Iguaçu IPRO had the highest RDM in the control application. The reduction in biomass is directly linked to ALS inhibition by nicosulfuron, which blocks the synthesis of branched-chain amino acids (leucine, isoleucine, and valine), compromising protein synthesis and consequently cell growth and biomass accumulation (Guo et al., 2023; Kumar et al., 2025).

Table 3 presents the transient chlorophyll a fluorescence parameters of soybean varieties subjected to nicosulfuron application at two times prior to sowing. These parameters are: Photosystem II quantum efficiency (Fv/Fm), electron transport quantum yield (Phi_Eo), dissipation energy quantum yield (Phi_Do), absorption-based performance index (Pi_Abs), absorbed energy flux per active reaction center (ABS/RC), energy flux trapped by active PSII (TRo/RC), electron transport flux per reaction center (ETo/RC), and dissipated energy flux (as heat and fluorescence) by active PSII (DIo/RC).

Table 3
Photosystem II quantum efficiency (Fv/Fm), Electron transport quantum yield (Phi_Eo), Dissipation energy quantum yield (Phi_Do), Absorption-based performance index (Pi_Abs), Energy flux absorbed per active reaction center (ABS/RC), Energy flux trapped by active PSII (TRo/RC), Energy flux absorbed per reaction center (ETo/RC), Energy flux dissipated (as heat and fluorescence) by active PSII (DLo/RC) of soybean varieties subjected to nicosulfuron application at two times prior to sowing

Varieties	Fv/Fm			Phi_Eo
	28 DBS	7 DBS	Control	28 DBS	7 DBS	Control
Guepardo IPRO	0.80 Aa	0.79 Aab	0.78 Aa	0.45 Ab	0.46 Aab	0.46 Aa
Olimpo IPRO	0.81 Aa	0.78 ABab	0.77 Ba	0.49 Aab	0.44 Bab	0.45 Aba
Tormenta CE	0.82 Aa	0.82 Aa	0.75 Ba	0.50 Aab	0.49 Aa	0.46 Aa
Iguaçu IPRO	0.81 Aa	0.82 Aa	0.78 Aa	0.52 Aa	0.49 ABa	0.45 Ba
M7601 i2x	0.80 Aa	0.75 Bb	0.79 Aa	0.49 Aab	0.43 Bb	0.47 Aba
ST700 i2x	0.81 Aa	0.79 Aab	0.79 Aa	0.49 Aab	046 Aab	0.46 Aa
CV (%)	2.50			5.67
LSD Column	0.04			0.05
LSD Line	0.03			0.04

	Phi_Do			Pi_Abs
	28 DBS	7 DBS	Control	28 DBS	7 DBS	Control
Guepardo IPRO	0.19 Aa	0.20 Aab	0.21 Aa	2.55 Aa	2.53 Aab	2.37 Aa
Olimpo IPRO	0.18 Aa	0.21 Aab	0.22 Aa	3.28 Aa	2.35 ABab	2.24 Ba
Tormenta CE	0.17 Ba	0.17 Bb	0.24 Aa	3.62 Aa	3.50 Aa	2.48 Ba
Iguaçu IPRO	0.20 Aa	0.18 Ab	0.21 Aa	3.58 Aa	3.52 Aa	2.47 Ba
M7601 i2x	0.19 Ba	0.24 Aa	0.20 Aba	2.96 Aa	1.86 Bb	2.50 Aba
ST700 i2x	0.18 Aa	0.20 Aab	0.20 Aa	3.16 Aa	2.66 Aab	2.64 Aa
CV (%)	11.00			20.83
LSD Column	0.02			1.21
LSD Line	0.03			0.99

	ABS/RC			TRo/RC
	28 DBS	7 DBS	Control	28 DBS	7 DBS	Control
Guepardo IPRO	2.26 Aa	2.11 Aa	2.26 Aa	1.82 Aa	1.67 Aa	1.78 Aa
Olimpo IPRO	2.06 Aa	2.19 Aa	2.29 Aa	1.67 Aab	1.72 Aa	1.77 Aa
Tormenta CE	2.03 Aa	2.02 Aa	2.14 Aa	1.67 Aab	1.66 Aa	1.60 Aa
Iguaçu IPRO	2.01 Aa	2.01 Aa	2.14 Aa	1.54 Ab	1.64 Aa	1.69 Aa
M7601 i2x	2.21 Aa	2.24 Aa	2.27 Aa	1.78 Aa	1.69 Aa	1.80 Aa
ST700 i2x	2.15 Aa	2.11 Aa	2.11 Aa	1.74 Aab	1.67 Aa	1.66 Aa
CV (%)	7.36			6.65
LSD Column	0.33			0.23
LSD Line	0.26			0.19

	ETo/RC			DIo/RC
	28 DBS	7 DBS	Control	28 DBS	7 DBS	Control
Guepardo IPRO	1.03 Aa	0.97 Aa	1.04 Aa	0.43 Aa	0.43 Aab	0.48 Aa
Olimpo IPRO	1.02 Aa	0.98 Aa	1.05 Aa	0.39 Ba	0.47 ABab	0.51 Aa
Tormenta CE	1.02 Aa	0.91 Aa	1.00 Aa	0.36 Ba	0.35 Bb	0.54 Aa
Iguaçu IPRO	0.97 Aa	0.99 Aa	0.97 Aa	0.37 Aa	0.36 Ab	0.45 Aa
M7601 i2x	1.08 Aa	0.97 Aa	1.07 Aa	0.42 Ba	0.55 Aa	0.47 Aba
ST700 i2x	1.05 Aa	0.98 Aa	1.15 Aa	0.40 Aa	0.43 Aab	0.44 Aa
CV (%)	10.52			15.75
LSD Column	0.22			0.14
LSD Line	0.18			0.11

• * Means followed by different lowercase letters in the Column and uppercase letters in the Row differ from each other by Tukey's test (p≤0.05). CV: Coefficient of variation; DBS: Days before sowing.

The maximum quantum efficiency of PSII (Fv/Fm) remained close to 0.80 for most varieties, constituting a typical value for unstressed plants. However, the M7601 i2x variety showed a significant reduction in Fv/ Fm when the herbicide was applied at 7 DBS, indicating possible photoinhibitory damage or stress in PSII during this shorter interval. On the other hand, varieties such as Tormenta CE and Iguaçu IPRO maintained high Fv/ Fm regardless of application, demonstrating resilience. Fv/Fm reflects the energy capture efficiency by open reaction centers; its reduction suggests an increase in the dissipation of non-photosynthetic energy (Akhter et al., 2021).

The quantum yield of electron transport (Phi_Eo) was reduced in the Olimpo IPRO and M7601 i2x varieties after application at 7 DBS, indicating damage to the electron transport chain, possibly due to thylakoid damage or induced oxidative stress (Bashir et al., 2021). However, the Iguaçu IPRO variety responded with an increase in Phi_Eo under herbicide application. The quantum yield of dissipation (Phi_Do) increased in the Tormenta CE control variety, suggesting activation of photoprotective mechanisms to dissipate excess energy (Ayyaz et al., 2020; Didaran et al., 2024).

The absorption-based performance index (Pi_Abs), which reflects integrated photosynthetic vitality, was superior for the Tormenta CE and Iguaçu IPRO varieties under herbicide application, indicating greater physiological tolerance and efficiency in the use of absorbed energy (Ghaffar et al., 2023). The absorbed energy flux (ABS/RC) did not differ between treatments regarding energy fluxes per reaction center, indicating that the structure of the light-harvesting antennae was not compromised (Mao et al., 2023).

The trapped energy flux (TRo/RC) did not vary with application within varieties, but Guepardo IPRO and M7601 i2x showed higher values than Iguaçu IPRO in the comparison between varieties at 28 DBS, suggesting a lower energy conversion capacity in the latter at that time (Khatri, Rathore, 2019; Mihaljević et al., 2021). The electron transport flux (ETo/RC) was not affected, showing that electron transport beyond the QA acceptor remained efficient. The dissipated energy flux (DIo/RC) was reduced by the herbicide application in the Olimpo IPRO (28 DBS) and Tormenta CE (both times) varieties, indicating less energy loss due to dissipation, which may be associated with greater photochemical efficiency under stress under these conditions. Finally, the M7601 i2x variant had a higher DIo/RC at 7 DBS, suggesting greater activation of dissipation mechanisms at this specific time (Zavafer; Mancilla, 2021).

3.2 Field assay

Table 4 presents the results for initial stand, plant height (initial and final), chlorophyll a and b, number of pods and grains per plant, thousand-grain weight, and yield of the Olimpo IPRO and Iguaçu IPRO varieties. As can be observed, no significant effects were observed for the interaction between variety and application time for these variables, nor for the application time factor alone. A significant effect was only observed for the variety factor.

Table 4
Means considering variety effects for initial and final stand, initial and final height, chlorophyll a and b, number of pods (NPP) and grains (NGP) per plant, thousand-grain weight (TGW) and yield variables of the Olimpo IPRO and Iguaçu IPRO varieties subjected to different nicosulfuron application intervals prior to sowing

Variables	Means: Varieties		CV (%)		LSD Varieties
	Olimpo IPRO	Iguaçu IPRO	Period	Variety
Initial stand (plants m-1)	12.07 b	15.41 a	5.69	5.81	0.6721
Initial height (cm)	8.74 b	10.07 a	5.01	7.81	0.3127
Final height (cm)	113.96 a	93.33 b	3.91	5.07	10.6199
Chlorophyll a	21.89 a	20.98 b	2.29	3.2	0.3254
Chlorophyll b	7.04 a	6.03 b	5.19	3.16	1.2534
NPP	60.88 a	47.36 b	24.1	13.59	8.6577
NGP	140.85 a	98.67 b	22.43	10.54	17.8304
TGW (g)	184.60 a	146.22 b	4.8	6.07	8.4587
Yield (kg ha-1)	4,625.08 a	3,986.38 b	9.94	12.81	514.4639

• CV (%): Coefficient of variation. Means with different letters on the same line differ from each other by the Student's t-test (LSD) (p≤0.05).

In turn, there was no significant effect of the variety x application time interaction in the field experiment, nor a main effect of the nicosulfuron application times on the analyzed variables (Tables 4 and 5). This indicates that the interval between herbicide application and sowing (7 to 28 DBS) did not negatively influence the establishment and final development of the plants under the study conditions.

Table 5
Mean final stand and stem diameter of the Iguaçu IPRO and Olimpo IPRO soybean varieties subjected to nicosulfuron application at four times prior to sowing

Application period	Final stand		Stem diameter
	Olimpo IPRO	Iguaçu IPRO	Olimpo IPRO	Iguaçu IPRO
28 DBS	12.33 Ba	14.69 Aa	3.88 Aa	3.44 Aba
21 DBS	11.67 Bab	14.73 Aa	3.83 Aa	3.28 Ba
14 DBS	11.62 Bab	14.44 Aa	3.75 Aa	3.52 Aba
7 DBS	10.87 Bb	13.23 Ab	3.85 Aa	3.12 Ba
Control	12.02 Bab	15.48 Aa	4.13 Aa	3.82 Aa
CV 1 (%)	3.12		11.11
CV 2 (%)	7.55		10.87
LSD Column	2.29		10.43
LSD Line	0.98		0.46

• Different lowercase means in the Column and uppercase means in the Row differ from each other by the Student's t-test (LSD) (p≤0.05); CV1: coefficient of variation of range; CV2: coefficient of variation of application times. DBS: days before sowing.

However, significant differences were observed between the Olimpo IPRO and Iguaçu IPRO varieties (Table 4). The Olimpo IPRO variety outperformed Iguaçu IPRO in final height, chlorophyll a and b content, number of pods per plant (NVP), number of grains per plant (NGP), thousand-grain weight (TGW), and yield. These characteristics are interrelated: greater height and higher chlorophyll content confer greater light interception capacity and photosynthetic efficiency, favouring photoassimilate accumulation and their allocation to reproductive structures, resulting in a greater number of grains and greater grain weight, culminating in the superior yield of Olimpo IPRO (Miglani et al., 2021; Simkin et al., 2022). Chlorophyll a acts directly in light energy conversion, while chlorophyll b broadens the absorption spectrum (Tanaka and Ito, 2025).

On the other hand, the Iguaçu IPRO variety showed higher initial stand and initial height values. The higher initial stand was expected due to the specific technical recommendation of higher sowing density for this variety. The greater initial height can be attributed to its earlier maturation (GM 6.7) compared to Olimpo IPRO (GM 7.7), which is generally associated with faster initial development.

Analysis of the final stand (Table 5) demonstrated that nicosulfuron application 7 DBS resulted in a significant reduction in the number of plants per meter in both varieties. This effect suggests the occurrence of deleterious effects on soybeans due to herbicide residue during crop emergence in scenarios where the interval between application and sowing is reduced, potentially impacting seed germination or seedling vigor (Bourdineaud, 2022).

Lastly, the Olimpo IPRO variety showed no differences between treatments and the control regarding stem diameter, demonstrating tolerance to herbicide residues. However, the Iguaçu IPRO variety exhibited significantly thinner stems when the herbicide was applied at 21 and 7 DBS compared to the control. A reduction in stem diameter can compromise the plant's structural stability, increasing susceptibility to lodging and potentially negatively affecting final yield (Xu et al., 2020). This result corroborates the greater sensitivity of the Iguaçu IPRO variety observed in some parameters in the greenhouse assay.

3.3 Similarity of varietal responses to nicosulfuron under different experimental conditions

The results obtained in the greenhouse and in the field show high consistency, indicating that the effects of nicosulfuron on soybeans are predominantly determined by varietal sensitivity, regardless of the evaluation environment. This convergence of results reinforces the robustness of the data and suggests that the physiological mechanisms associated with tolerance or sensitivity to nicosulfuron are intrinsic to the genotype, manifesting themselves similarly under controlled conditions and under environmental variability. Furthermore, regardless of the experimental condition, the results demonstrated that nicosulfuron has a deleterious effect depending on its positioning, which indicates that the residual effect of this herbicide can act consistently on fundamental morphophysiological processes for soybean plants. More objectively, when the interval between herbicide application and soybean sowing is not respected, greater negative impacts were observed on the response variables analyzed in soybeans.

Despite the initial sensitivity observed in some cultivars, the field results showed that this does not always translate into proportional losses in final yield. The Olimpo IPRO cultivar, for example, even though it showed greater initial sensitivity in the greenhouse, stood out in the field in characteristics directly related to productivity. This result indicates the existence of compensatory mechanisms throughout the cycle, such as greater efficiency in vegetative recovery or better reproductive capacity, which are not fully captured in initial evaluations in a controlled environment. In general, the similarity between the experiments reinforces that varietal sensitivity to nicosulfuron is a key factor for the safe management of this herbicide in succession systems with soybeans. The integration of the results shows that greenhouse trials are effective in identifying tolerance patterns, while field studies complement this information by revealing the recovery capacity of the cultivars and their effects on productivity. Thus, the choice of more tolerant cultivars, combined with respect for application intervals, is an essential strategy to mitigate phytotoxicity risks and ensure the productive potential of the crop.

4. Conclusions

The nicosulfuron herbicide application prior to soybean sowing had a distinct impact on the initial development of the evaluated varieties, highlighting variations in varietal sensitivity to herbicide residue in the soil. The Guepardo IPRO, ST700 i2x, and Tormenta CE varieties stood out for showing greater tolerance.

On the other hand, the M7601 i2x, Olimpo IPRO, and Iguaçu IPRO varieties showed greater sensitivity to nicosulfuron, with persistent reduction in plant height, biomass accumulation, and root development, in addition to physiological changes in some cases.

Although the timing of nicosulfuron herbicide application in the field assay did not significantly influence most of the variables analysed, the choice of variety directly impacted the agronomic performance of soybeans. The Olimpo IPRO variety stood out with the best results in characteristics related to yield, such as final height, chlorophyll content, number of pods and grains, TGW, and final yield. The Iguaçu IPRO variety showed higher values for stand and initial height, with these characteristics being influenced by its earlier cycle and denser sowing stand. Choosing more tolerant varieties and respecting the application interval are fundamental to minimizing phytotoxic effects and preserving the productive potential of soybeans.

The results of this study are valuable and allow us to understand how factors such as soybean genotype and the interval between nicosulfuron application and soybean sowing can impact crop development. Further studies are important, as they will allow for a better understanding of soybean cultivation in environments with varying soil classes, as well as the application of different doses of nicosulfuron.

Acknowledgements

The authors thank the Instituto Federal Goiano (IFGoiano) for its institutional support and the infrastructure provided for conducting the research. We also thank the Centro Tecnológico COMIGO (CTC) for providing materials and experimental conditions essential to the development of the study. We acknowledge the support of the IF Goiano Weed Laboratory and the support of the CNPq responsible for granting the scholarship.

• Funding
  This study was funded by the National Council for Scientific and Technological Development (CNPq), through the Academic Master's and Doctoral Program for Innovation - MAI/DAI (Process No. 403524/2020-2) and by Centro Tecnológico COMIGO (CTC), and by Centro Tecnológico COMIGO (CTC) and Instituto Federal Goiano (Process No. 23218.001766.2026-17).

Data Availability Statement

The data supporting the findings of this study are available from the corresponding authors upon reasonable request.

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Edited by

• Editor in Chief:
  Anderson Nunes Gabardo

• Associate Editor:
  Anderson Nunes Gabardo

Publication Dates

• Publication in this collection
  31 Aug 2026
• Date of issue
  2026

History

• Received
  26 Jan 2026
• Accepted
  26 May 2026