© The Authors, 2025, Published by the Universidad del Zulia*Corresponding author: jahuerta@uach.mx
Keywords:
Humic substance
Legume
Soilless
Biostimulants
Eect of fulvic acid on the growth of hydroponic pea (Pisum sativum L.) microgreens
Efecto del ácido fúlvico en el crecimiento de microvegetales hidropónicos de guisante (Pisum
sativum L.)
Efeito do ácido fúlvico no crescimento de microgreens de ervilha hidropônica (Pisum sativum L.)
Aldo Gutiérrez Chávez
Martha Irma Balandrán Valladarez
Rosa María Yáñez Muñoz
Jared Hernández Huerta*
Rev. Fac. Agron. (LUZ). 2025, 42(3): e254232
ISSN 2477-9407
DOI: https://doi.org/10.47280/RevFacAgron(LUZ).v42.n3.III
Crop production
Associate editor: Dra. Evelyn Pérez Pérez
University of Zulia, Faculty of Agronomy
Bolivarian Republic of Venezuela
Autonomous University of Chihuahua, Faculty of
Agrotechnological Sciences, Pascual Orozco, Chihuahua,
31350, C.P. 31000, Chih, Mexico.
Received: 13-04-2025
Accepted: 02-06-2025
Published: 30-06-2025
Abstract
Fulvic acid is a widely recognized biostimulant due to its
benets in traditional crops; however, its application in hydroponic
systems, particularly in microgreen production, is not well
documented. This study evaluated the eect of fulvic acid on the
growth of hydroponic pea microgreens (Pisum sativum L.). The
experimental design was completely randomized and consisted of
four treatments (n=5): nutrient solution (NS), fulvic acid solution
0.01 % (FA), NS + FA, and water (control). After 12 days, growth
and biochemical parameters were measured. The results showed that
NS and NS+FA treatments signicantly increased stem length (7.73
cm and 7.28 cm), fresh weight (0.613 g and 0.618 g), and yield (6.15
kg.m
-2
) compared to the FA treatment or control. The FA treatment
increased stem diameter (2.38 mm) but did not signicantly
increase biomass. Biochemical analysis showed that FA and control
had higher nitrate content, while NS and NS+FA reduced nitrate
accumulation. Antioxidant capacity, chlorophyll content, and color
index were similar among treatments. However, the pH increased
with the application of fulvic acid. Fulvic acid alone moderately
improved growth but was less eective than the nutrient solution.
The combination of fulvic acid with a complete nutrient solution
did not produce additive eects, highlighting the importance of
balanced nutrition in hydroponic microgreen production.
This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.
Rev. Fac. Agron. (LUZ). 2025, 42(3): e254232 July-September. ISSN 2477-9409.
2-6 |
Resumen
El ácido fúlvico es un bioestimulante reconocido por sus benecios
en cultivos tradicionales; sin embargo, su aplicación en sistemas
hidropónicos, particularmente en la producción de microvegetales,
no está bien documentada. Este estudio evaluó el efecto del ácido
fúlvico sobre el crecimiento de microvegetales de chícharo (Pisum
sativum L.) cultivados en hidroponía. El diseño experimental fue
completamente al azar y consistió en cuatro tratamientos (n=5):
solución nutritiva (SN), solución de ácido fúlvico al 0,01 % (AF),
SN+AF y agua (control). Después de 12 días, se midieron parámetros
de crecimiento y bioquímicos. Los resultados mostraron que los
tratamientos SN y SN+AF incrementaron signicativamente la
longitud del tallo (7,73 cm y 7,28 cm), el peso fresco (0,613 g y 0,618
g) y el rendimiento (6,15 kg.m
-2
) en comparación con AF o el control.
El tratamiento AF incrementó el díametro de tallo (2,38 mm), pero
no aumentó signicativamente la biomasa. El análisis bioquímico
mostró que el AF y el control presentaron un mayor contenido de
nitratos, mientras que los tratamientos SN y SN+AF redujeron la
acumulación de estos. La capacidad antioxidante, el contenido de
clorola y el índice de color fueron similares entre tratamientos.
Sin embargo, el pH aumentó con la aplicación de ácido fúlvico. El
ácido fúlvico mejoró moderadamente el crecimiento, pero fue menos
efectivo que la solución nutritiva. La combinación de ácido fúlvico
con una solución nutritiva completa no produjo efectos aditivos,
lo que resalta la importancia de una nutrición equilibrada en la
producción hidropónica de microvegetales.
Palabras clave: sustancia húmica, leguminosa, cultivo sin suelo,
bioestimulantes.
Resumo
O ácido fúlvico é um bioestimulante reconhecido por seus
benefícios em culturas tradicionais; Entretanto, sua aplicação em
sistemas hidropônicos, particularmente na produção de microgreens,
não está bem documentada. Este estudo avaliou o efeito do ácido
fúlvico no crescimento de microgreens de ervilha (Pisum sativum
L.) cultivados hidroponicamente. O delineamento experimental
foi inteiramente casualizado e consistiu em quatro tratamentos
(n=5): solução nutritiva (SN), solução de ácido fúlvico 0,01 %
(AF), SN+AF e água (controle). Após 12 dias, foram medidos os
parâmetros de crescimento e bioquímicos. Os resultados mostraram
que os tratamentos SN e SN+AF aumentaram signicativamente o
comprimento do caule (7,73 cm e 7,28 cm), o peso fresco (0,613 g
e 0,618 g) e o rendimento (6,15 kg.m
-2
) em comparação ao AF ou
ao controle. O tratamento AF aumentou o diâmetro do caule (2,38
mm), mas não aumentou signicativamente a biomassa. A análise
bioquímica mostrou que AF e controle apresentaram maior teor de
nitrato, enquanto os tratamentos SN e SN+AF reduziram o acúmulo
de nitrato. A capacidade antioxidante, o teor de clorola e o índice
de cor foram semelhantes entre os tratamentos. Entretanto, o pH
aumentou com a aplicação de ácido fúlvico. O ácido fúlvico melhorou
moderadamente o crescimento, mas foi menos ecaz que a solução
nutritiva. A combinação de ácido fúlvico com uma solução nutritiva
completa não produziu efeitos aditivos, destacando a importância da
nutrição balanceada na produção de microgreens hidropônicos.
Palavras-chave: substância húmica, leguminosa, cultivo sem solo,
bioestimulantes
Introduction
The production of microgreens has become increasingly important
in recent years due to their high nutritional content, short cultivation
cycle, and growing demand in gourmet and functional food markets
(Choe et al., 2018; Rouphael et al., 2021). These small vegetables
harvested at early developmental stages are rich in vitamins, minerals,
antioxidants, and bioactive compounds, making them attractive to
consumers interested in healthy and sustainable foods (Sharma et al.,
2022; Xiao et al., 2012). Among the species grown as microgreens,
pea (Pisum Sativum L.) stands out for its nutritional prole, high
protein content, and culinary versatility, justifying the exploration of
strategies to optimize its growth and quality in hydroponic systems
(Xiao et al., 2019; Ebert, 2022).
Biostimulants, such as fulvic acid, have emerged as an innovative
practice to enhance agricultural production (Canellas et al., 2015;
Bell et al., 2022). Fulvic acids, soluble fractions of organic matter,
possess unique properties that positively aect plant metabolism
by enhancing nutrient uptake, improving photosynthetic eciency,
and increasing tolerance to abiotic stress (Canellas et al., 2015;
Hasanuzzaman et al., 2021; Mosaad et al., 2024). Although their
ecacy has been extensively studied in traditional crops, their
application in microgreens, especially in hydroponic systems,
requires further research to fully understand their eects on growth
and quality parameters (Drobek et al., 2019; Sharma et al., 2022).
In the case of pea microgreens, harvesting is typically recommended
between 10 and 14 days after germination, when the shoots reach a
height of 7 to 10 cm, and exhibit an intense green color, which are
considered key quality indices for market acceptance (Tallei et al.,
2024). These morphological characteristics are critical in determining
harvest timing and consumer preference, as they are directly related to
visual appeal, texture, and nutritional content. Therefore, optimization
of growth conditions and inputs such as biostimulants is essential to
meet quality standards and improve yield consistency in commercial
production.
Previous studies have shown that biostimulants can induce
signicant changes in plant development through physiological and
biochemical mechanisms, such as increased chlorophyll production,
enzymatic activity, the accumulation of bioactive compounds
(Graziani et al., 2022; Sharma et al., 2022; Anastacio-Angel et al.,
2024).
Howeverplant response to fulvic acid can vary depending on
the cultivated species and production system conditions (Zhang
et al., 2021). In this context, it is important to evaluate how fulvic
acid aects the growth and yield of pea microgreens in hydroponic
systems, considering the growing need for sustainable and ecient
agricultural practices.
This aim of this study was to evaluate the eect of applied fulvic
acid, on the growth, biochemical composition, and yield of pea tendril
microgreens grown hydroponically, in order to determine its potential
as a biostimulant for sustainable microgreen production. This work
will contribute to the understanding of the potential benets of fulvic
acid in sustainable production systems and provide a scientic basis
for its application in urban agriculture and functional food production.
This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.
Gutierrez et al. Rev. Fac. Agron. (LUZ). 2025, 42(3): e254232
3-6 |
Materials and methods
Localization experiment
The experiment was conducted at the Applied Microbiology,
Plant Pathology, and Post-harvest Physiology Laboratory of the
Autonomous University of Chihuahua, Chihuahua, MX (28°39’24’’
N, 106°05’12’’ W) during November and December 2024.
Plant and fulvic acid material
Organic tendril pea microgreens (Pisum sativum L.) seeds
(Johnny’s Selected Seeds, USA) were used for the test. An aqueous
solution of fulvic acids derived from leonardite (K-Tionic®, Arysta
LifeSience México, MX) containing 25 % organic fulvic acid.
Experimental setup
Pea seeds were washed twice with tap water, soaked in a 0.12 %
H
2
O
2
solution for 6 h, drained and placed directly in polystyrene trays
(13x13x8 cm) (S-22911, Uline México, MX) containing a plastic
mesh (1.8 mm) (B0BXKV98MF, Spkaodngo, USA) 2 cm above the
bottom, without substrate, at 1 seed per cm
2
(Verlinden, 2020), and
placed in the dark at 24 °C. After germination, two-day-old tendril
pea plants were transferred to a growth chamber with a photoperiod
of 16 h light/8 h dark at 28 °C/ 18 °C, 3,500 lux LED light (Goodwill
az-energy®, 20460, MX), and 70 ± 2 % relative humidity. The
microgreens were watered every two days with dierent solutions:
A) Steiner nutrient solution (NS) composed of (ppm): 126 NO
3
-
, 42
NH
4
+
, 31 PO
4
3-
, 274 K
+
, 181 Ca
2+
, 48.6 Mg
2+
, 112 SO
4
2-
, 1.3 Fe-EDTA,
0.8 Mn-EDTA, 0.3 Zn-EDTA, 0.06 Cu-EDTA, 0.4 B, and 0.06 Mo
(pH 6.0, EC 2.3 mS.cm
-1
); B) Fulvic acid solution (0.01 %, pH 6.0,
EC 0.45 mS.cm
-1
), based on the dosage recommended in the technical
data sheet of the product; C) NS + FA (pH 6.0, EC 2.5 mS.cm
-1
); and
D) destilled water.
Parameters evaluated
The growth and biochemical parameters of the microgreens were
evaluated on day 12 after sowing.
Growth parameters
Ten seedlings from each replicate were cut at the collar region.
Stem length and diameter were measured using a digital caliper
(Starret®, EC799A-6/150, USA). Stipular leaf area was determined
using ImageJ 1.46r software. The number of tendrils was recorded,
and fresh weight (FW) and dry weight (DW) were measured using
an analytical balance (XT-220A, Precisa Instruments®, Switzerland)
after drying at 60 °C for 48 h, in a forced-air convection oven (SMO3,
Shel Lab®, USA). The water content (WC) of the microgreen
seedlings was determined using the following equation (Eq. 1):
(Eq. 1)
Yield: The yield of pea microgreens was calculated based on a
seeding density of 1 seed per cm
2
, using the following equation (Eq. 2):
Yield (kg.m
2
)= Fresh weight of seedlings (kg) x seedlings per m
2
(Eq. 2)
Biochemical parameters
pH: 5 g of seedlings were macerated, and the pH was measured
using a pH meter (Checher® pH Tester HI98103, Hanna Instruments,
USA). Total soluble solids (TSS) were expressed in °Brix: A drop of
microgreen juice was placed on a digital refractometer (Automatic
Refractometer Smart-1, Japan).
Color index (CI): Color was measured using the CIE L*a*b*
system with a digital colorimeter (Minolta Chroma Meter CR-310;
Konica Minolta Optics, Japan). The CI was calculated with the
following equation (Eq. 3):
CI= (1000 * a) / (L * b) (Eq. 3)
Photosynthetic pigment content
A 0.1 g sample of fresh leaves was macerated with 4 mL of 80 %
acetone (v/v) and centrifuged at 3,000 rpm for 5 min. The supernatant
was measured at 663, 470, and 645 nm using a UV spectrophotometer
(Model 60S Evolution, Thermo Scientic, USA) (Lichtenthaler &
Wellburn, 1983). Pigment concentrations were calculated as follows
(Eq 4, 5 and 6):
Where: V= volume (mL) of 80 % acetone, W is the fresh weight
(FW) of the sample (g).
Antioxidant activity
Fresh samples (10 g) were homogenized with 20 mL of 80 %
ethanol and diluted to 100 mL with destilled water. The mixture
was stirred for 10 min, ltered (Whatman No. 1), and 0.1 mL of
the extract was mixed with 3.9 mL of 2,2-diphenyl
-1
-picrylhydrazyl
(DPPH; 0.025 g.L
-1
) ethanolic solution. After 60 min in the dark
at 25 °C, the absorbance was measured at 515 nm using a UV-vis
spectrophotometer. The results were expressed as the percentage of
DPPH radical inhibition, calculated using the following equation (Eq.
7) (Rodríguez-Roque et al., 2013):
Where: C
abs
=absorbance of the control, SM
abs
= absorbance of the
sample extract.
Nitrate content: A 1 g sample of fresh leaves was homogenized
in 3 mL of distilled water, centrifuged at 4,000 rpm for 15 min, and
20 μL of the supernatant was mixed with 80 μL of 5 % sulfuric acid-
salicylic acid and 3 mL of 1.5 N NaOH. After 10 min, the absorbance
was measured at 410 nm using a UV-visible spectrophotometer, and
the nitrate concentration was calculated using a KNO
3
standard curve
(0, 1, 2.5, 5, 7.5, 10 mM; R
2
= 0.995) (Toscano et al., 2021).
Statistical analysis
The experiment was set up as a completely randomized design with
four treatments: FA, NS + FA, NS, and puried water (control), each
replicated ve times. Growth and biochemical data were subjected to
Shapiro-Wilk and Levene tests for normality and homoscedasticity.
Depending on these results, data were analyzed by analysis of
variance (ANOVA) with the Tukey test or non-parametric Kruskal-
Wallis with Dunn test (p<0.05). A Principal Component Analysis
(PCA) was performed on key variables to evaluate the inuence of
fulvic acid on pea microgreens, validated by Bartlett′s test (p<0.01)
and Kaiser-Meyer-Olkin (KMO) measure (>0.60). Data analysis was
performed with Jamovi software 2.5.2.0.
Results and discussion
Growth parameters
The application of dierent treatments signicantly aected the
growth of tendril pea microgreens (table 1).

(
%
)
=
 ()()
()
100 1
(Eq.4)
(Eq.5)
(Eq.6)
DPPH inhibition (%) =
C
abs
- SM
abs
C
abs
100 1
(Eq.7)
Chlorophyll a
(
mg.g
-1
FW
)
= (12.21 × A
663
-2.81 × A
645
) × V (1000 ×W)
1
Chlorophyll b
(
mg.g
-1
FW
)
= (20.13 × A
645
-5.03 × A
663
)× V (1000 × W)
1
Carotenoids
(
mg.g
-1
FW
)
=
(
1000 × A
470
-3.27 × Chl
a
-104 × Chl
b
)
229
× V/(1000 × W) 1
This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.
Rev. Fac. Agron. (LUZ). 2025, 42(3): e254232 July-September. ISSN 2477-9409.
4-6 |
Table 1. Growth parameters of tendril pea microgreens treated
with fulvic acid cultivated in a hydroponic system for
12 days.
Parameters
Treatments
CV
Control FA NS NS + FA
Stem length (cm)
2
3.70
c
4.65
b
7.73
a
7.28
a
18.04
Stem diameter (mm)
1
2.34
ab
2.38
a
2.17
c
2.26
b
7.53
Tendrils Number
2
4.32
a
3.76
b
4.22
a
4.36
a
12.59
Leaf area stipulate
(cm
2
)
2
3.08
c
3.63
b
5.40
a
5.35
a
18.11
Fresh shoot weight (g)
2
0.225
c
0.333
b
0.613
a
0.618
a
17.84
Dry shoot weight(g)
2
0.028
c
0.040
b
0.059
a
0.061
a
17.14
Water content (%)
2
86.88
b
87.48
b
90.08
a
89.84
a
3.43
Dierent superscript letters in the same row indicate signicant dierences according to the
Games-Howell test
1
or Dunn test
2
at the 0.05 level. Control= destillated water, FA= fulvic acid,
NS= Nutrient solution, CV= coecient of variation.
For stem length, the NS and NS+FA treatments showed statistically
higher values (7.73 cm and 7.28 cm, respectively) compared to FA
(4.65 cm) and control (3.70 cm). These results indicate that nutrient
supplementation, with or without fulvic acid, enhances elongation,
probably due to the increased availability of essential ions such
as K
+
and NO₃⁻, which are known to promote cell expansion and
division (Sano et al., 2009). However, the addition of fulvic acid to
the nutrient solution (NS+FA) did not further increase stem length
compared to NS alone, suggesting no synergistic eect, which may
be due to altered solubility or nutrient uptake interactions, as noted by
Wang et al. (2022).
In terms of stem diameter, the FA treatment (2.38 mm) was
statistically superior to NS (2.17 mm), while NS+FA (2.26 mm)
showed intermediate values. This may reect the gibberellic acid-
like eects of fulvic acid that promote secondary growth (Pizzeghello
et al., 2001), although the NS alone appeared to be less eective,
possibly due to the prioritization of elongation over thickening in
nitrogen-rich environments.
The number of tendrils was signicantly reduced in FA compared
to the other treatments. NS, NS+FA, and control showed no signicant
dierences, with an average of 4.3 tendrils per plant. This suggests
that fulvic acid alone may dierentially aect morphogenetic
patterns, possibly tangentially by modulating the auxin-cytokinin
ratio (Muscolo et al., 2007). Stipular leaf area increased signicantly
in all treatments compared to the control, with the highest values in
NS and NS+FA (5.40 and 5.35 cm
2
, respectively) and a moderate
increase in FA (3.36 cm
2
). These improvements could be related to
enhanced nitrogen assimilation and carbon metabolism, as suggested
by He et al. (2021), especially in nutrient-enriched systems.
For fresh and dry shoot weight, NS and NS+FA achieved the
highest biomass values (0.613 g and 0.618 g fresh; 0.059 g and
0.061 g dry, respectively), signicantly higher than FA and control.
While FA alone improved biomass compared to control, its eect
was inferior to NS-based treatments. These results suggest that the
combination of macro-and micronutrients is more critical for biomass
accumulation than fulvic acid alone, although FA may still contribute
to early-stage development.
Water content was not signicantly dierent among treatments,
although NS and NS+FA presented slightly higher averages (~90 %)
than FA and control (~87 %). This indicates that fulvic acid did not
adversely aect water retention and that the high water content in
nutrient-treated seedlings reects better turgor and hydration under
optimal mineral nutrition.
Overall, these results show that while fulvic acid alone improves
some growth parameters compared to the control, the nutrient solution
treatments (NS+FA) provide the most substantial benets. The lack of
additive eects in NS+FA may be related to chemical interactions that
limit FA availability or action.
Yield
The yield of tendril pea microgreens was signicantly aected by
the treatments (gure 1a).
Figure 1. Yield of tendril pea microgreens treated with fulvic acid
(FA) cultivated in a hydroponic system for 12 days.
Control= destillate water, NS= nutrient solution. Bars
with same letters show no signicant dierences (p<0.05,
Tukey test).
The NS and NS+FA treatments achieved the highest yields, with
statistically similar values averaging 6.15 kg.m
-2
, indicating that the
application of Steiner nutrient solution either alone or in combination
with fulvic acid, enhanced the increased biomass production. In
contrast, the application of FA alone resulted in a moderate yield of 3.3
kg.m
-2
, an increase of 43.47 % over the control (2.3 kg.m
-2
), but was
signicantly lower than the NS-based treatments. This suggests that
while fulvic acid has a stimulatory eect on plant growth, probably
due to its role in improving nutrient uptake and metabolic activation
(Muscolo et al., 2007), it cannot match the contribution of a complete
nutrient formulation in supporting maximum biomass accumulation.
The NS+FA treatment did not signicantly exceed the
performance of NS alone, consistent with observations from the
morphological parameters. This lack of additive eect may be due
to chemical interactions that reduce the bioavailability or functional
eciency of fulvic acid in nutrient-rich environments (Wang et al.,
2022), or possibly to saturation eects where the nutrient solution
already meets or exceeds the nutritional needs of the plant.
These results support the idea that while fulvic acid can be
benecial, its most eective use may be in nutrient-limited systems
or in the early stages of growth, rather than as an additive to already
balanced nutrient solutions. Additionally, the yield levels observed
for the NS and NS+FA treatments are within the upper range of
microgreen productivity under hydroponic conditions reported in the
literature (Xiao et al., 2012), further validating the eectiveness of
the selected nutrient regime.
Visual quality dierences among treatments were evident (gure
1b). NS and NS+FA produced denser, more upright, and visually
stronger microgreens compared to the control and FA treatments,
which appeared sparser and shorter. These visual dierences are
consistent with the yield and morphological data and reect more
robust development under nutrient-enriched conditions. Minor
mechanical damage during harvest was observed in all treatments due
to dierences in plant height; however, this did not compromise the
overall appearance for market purposes.
This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.
Gutierrez et al. Rev. Fac. Agron. (LUZ). 2025, 42(3): e254232
5-6 |
Biochemical parameters
The biochemical prole of tendril pea microgreens was generally
not signicantly aected by the application of FA, except for pH,
which showed signicant variation among treatments (table 2).
Table 2. Biochemical parameters of tendril pea microgreens
treated with fulvic acid and grow in a hydroponic
system for 12 days.
Parameters
Treatments
Control FA NS NS + FA CV
pH 5.85
b
6.26
a
6.24
a
6.28
a
1.64
TSS 11.81
a
11.11
a
9.81
b
9.32
b
5.55
Color Index -21.51
a
-24.65
a
-24.95
a
-24.82
a
25.50
Chlorophyll a 1.27
a
1.26
a
1.26
a
1.026
a
0.93
Chlorophyll b 0.76
a
1.04
a
0.97
a
1.02
a
17.19
Carotenoids 0.61
a
0.64
a
0.63
a
0.64
a
3.25
Antioxidant Ca-
pacity
48.90
a
50.70
a
51.00
a
50.90
a
3.28
Nitrates 2,572.42
a
2,616.25
a
1,400.56
b
1,434.17
b
19.91
Dierent superscript letters in the same row indicate signicant dierences according to the
Tukey test at the 0.05 level. Control= destillate water, FA= fulvic acid, NS= Nutrient solution.
CV= coecient of variation.
The pH of the microgreens increased with the FA, NS, and NS+FA
treatments compared to the control, with values ranging from 6.24 to
6.28, signicantly higher than the control (5.85). This increase may
be attributed to the carboxylic and phenolic groups present in fulvic
acids, which can alter the rhizosphere pH and internal tissue chemistry
(Muscolo et al., 2007). However, no signicant dierences were
observed among FA, NS and NS+FA, suggesting that fulvic acids and
mineral nutrients may have overlapping eects on pH modulation.
Higher pH in plant tissues has been associated with improved sensory
quality and shelf life in microgreens, contributing to reduced acidity,
improved avor perception, and microbial stability (Tallei et al.,
2024). These factors are important for increasing market value and
consumer acceptance in commercial production (Seth et al., 2025).
Total soluble solids (TSS) were signicantly higher in the
control and FA treatments (11.81 and 11.11 ºBrix, respectively),
while NS and NS+FA treatments had lower values (9.81 and 9.31
ºBrix, respectively). This suggests that nutrient-rich environments
may dilute sugar concentrations due to increased vegetative growth.
The higher TSS in the control may also reect stress-related sugar
accumulation due to nutrient deciency, as noted by Lin et al. (2016).
No signicant dierences in color index, chlorophyll a, chlorophyll
b, or carotenoids were observed among treatments, indicating that
neither FA nor nutrient solutions signicantly aected pigment
synthesis. Although NS and NS+FA treatments showed a trend
toward higher chlorophyll b and carotenoid content, the variation was
not statistically signicant. These results are consistent with studies
suggesting that chlorophyll biosynthesis requires not only adequate
nitrogen but also light quality cues, which may have remained stable
among treatments (Gao et al., 2023).
Antioxidant capacity remained statistically similar across
treatments, averaging around 50 %, suggesting that FA and nutrients
did not stimulate secondary metabolite production under the given
conditions. This is consistent with the ndings of Márquez-García et
al. (2011), where limited abiotic stress did not activate antioxidant
pathways in hydroponically grown legumes.
A signicant dierence was observed in nitrate accumulation,
with the control and FA treatments showing signicantly higher
concentrations (2,572 and 2,616 mg.kg⁻¹, respectively) compared
to NS and NS+FA (1,400 and 1,434 mg.kg⁻¹, respectively). This
indicates a limited capacity for nitrate assimilation in the absence
of a complete nutrient prole. As noted by Nardi et al. (2002), the
conversion of nitrate to amino acids requires cofactors and energy
sources provided by a balanced nutrition, which were absent in the
control and FA-only treatments.
Principal Component Analysis (PCA) provided further insight
into treatment eects (gure 2).
Figure 2. Principal components analysis of tendril pea microgreens
treated with fulvic acid (FA) and grown in a hydroponic
system for 12 days. TSS= Total soluble solids, LAS= leaf
area stipular, WC= water content, SL=Stem length, TN=
tendril number, FW= fresh weight.
The rst two components (PC1 = 56.9 %, PC2 = 17.1 %) explained
74.0 % of the total variance. The control was associated with higher
nitrate and TSS levels, suggesting a biochemical prole typical of
nutrient-decient but metabolically stressed plants. The FA treatments
showed a weak association with chlorophyll a, indicating a limited
photosynthetic enhancement. In contrast, NS+FA was associated
with improvements in morphological traits such as stem length (SL),
tendril number (TN), fresh weight (FW), and water content (WC).
The NS treatment, although more dispersed in its responses, was
associated with chlorophyll b and carotenoids, suggesting a slight
advantage in pigment biosynthesis and light-harvesting eciency.
These results suggest that while fulvic acids alone may slightly
alter internal pH and nitrate metabolism, their combination with
nutrient solution does not improve biochemical characteristics beyond
what is achieved by nutrients alone. This highlights the importance
of nutrient completeness over biostimulant supplementation in
optimizing biochemical composition, especially under non-stressful
hydroponic conditions.
Conclusions
The application of fulvic acid at 0.01 % in pea tendril microgreens
grown in a hydroponic system showed a positive, although limited,
This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.
Rev. Fac. Agron. (LUZ). 2025, 42(3): e254232 July-September. ISSN 2477-9409.
6-6 |
eect, especially under nutrient-limited conditions. The FA treatment
improved morphological variables such as stem diameter by 25.6 %
and fresh weight by 48 % and dry weight by 42.8 % compared to the
control, and increased yield by 43 %, demonstrating its potential as a
biostimulant in systems without mineral fertilization.
However, when considering all variables were considered
simultaneously through multivariate analysis, FA did not outperform
the complete nutrient solution and showed no synergistic eect when
combined with it. Biochemically, its application was associated with
higher nitrate and soluble solids accumulation, without improvements
in pigments, antioxidant capacity, or visual quality.
These results suggest that fulvic acid can partially modulate growth
and metabolism during early development, but the nutrient context
strongly conditioned its ecacy. Therefore, fulvic acid represents a
viable alternative to stimulate microgreen development under limited
nutrient conditions or as a complementary strategy. However, it does
not replace the need for balanced mineral fertilization when the goal
is to maximize productivity, quality, and commercial consistency of
the crop.
Literature cited
Anastacio-Angel, G., González-Fuentes, J.A., Zermeño-González, A., Robledo-
Olivo, A., Lara-Reimers, E.A., & Peña-Ramos, F.M. (2024). Efecto
de bioestimulantes en crecimiento, siología y calidad bioquímica
de frambuesa (Rubus idaeus L.) sometida a estrés hídrico. Terra
Latinoamericana, 42,1-4. https://doi.org/10.28940/terra.v42i0.1772
Bell, J.C., Bound, S.A., & Buntain, M. (2022). Biostimulants in agricultural and
horticultural production, In I. Warrington (Ed.), Horticultural Reviews,
49, (pp. 35-95). https://doi.org/10.1002/9781119851981.ch2
Canellas, L.P., Olivares, F.L., Aguiar, N.O., Jones, D.L., Nebbioso, A., Mazzei,
P., & Piccolo, A. (2015). Humic and fulvic acids as biostimulants in
horticulture. Scientia Horticulturae, 196, 15-27. https://doi.org/10.1016/j.
scienta.2015.09.013
Choe, U., Yu, L.L., & Wang, T.T (2018). The science behind microgreens as an
exciting new food for the 21st century. Journal of Agricultural and Food
Chemistry, 60(31), 7644-7651. https://doi.org/10.1021/jf2057816
Drobek, M., Fraś, A., Molas, J., & Kurasiak-Popowska, D. (2019). Plant
biostimulants: Importance of the quality and yield of horticultural crops
and the improvement of biological and physiological processes. Folia
Horticulturae, 31(2), 139-149. https://doi.org/10.2478/fhort-2019-0012
Ebert, A.W. (2022). Sprouts and microgreens: Novel food sources for healthy
diets. Plants, 11(4), 571. https://doi.org/10.3390/plants11040571
Gao, Y., Chen, S., Y., & Shi, Y. (2023). Eect of nano-calcium carbonate on
morphology, antioxidant enzyme activity and photosynthetic parameters
of wheat (Triticum aestivum L.) seedlings. Chemical and Biological
Technologies in Agriculture, 10(1), 31. https://doi.org/10.1186/s40538-
023-00404-9
Graziani, G., Cirillo, A., Giannini, P., Conti, S., El-Nakhel, C., Rouphael, Y., &
Di Vaio, C. (2022). Biostimulants improve plant growth and bioactive
compounds of young olive trees under abiotic stress conditions.
Agriculture, 12(2), 227. https://doi.org/10.3390/agriculture12020227
Hasanuzzaman, M., Parvin, K., Bardhan, K., Nahar, K., Anee, T.I., Masud, A.A.C.,
& Fotopoulos, V. (2021). Biostimulants for the regulation of reactive
oxygen species metabolism in plants under abiotic stress. Cells, 10(10),
2537 https://doi.org/10.3390/cells10102537
He, X., Zhang, H., Ye, X., Hong, J., & Ding, G. (2021). Nitrogen assimilation
related genes in Brassica napus: Systematic characterization and
expression analysis identied hub genes in multiple nutrient stress
responses. Plants, 10(10), 2160. https://doi.org/10.3390/plants10102160
Lichtenthaler, H.K., & Wellburn, A.R. (1983). Determinations of total carotenoids
and chlorophylls a and b of leaf extracts in dierent solvents. Biochemical
Society Transactions, 11(5), 591-592. https://doi.org/10.1042/bst0110591
Lin, X.Y., Ye, Y.Q., Fan, S.K., Jin, C.W., & Zheng, S.J. (2016). Increased sucrose
accumulation regulates iron-deciency responses by promoting auxin
signaling in Arabidopsis plants. Plant Physiology, 170(2), 907-920.
https://doi.org/10.1104/pp.15.01598
Márquez-García, B., Horemans, N., Cuypers, A., Guisez, Y., & Córdoba, F. (2011).
Antioxidants in Erica andevalensis: A comparative study between wild
plant and cadmium-exposed plants under controlled conditions. Plant
Physiology and Biochemistry, 49(1), 110-115. https://doi.org/10.1016/j.
plaphy.2010.10.007
Mosaad, I.S., Selim, E.M.M., Gaafar, D.E., & Al-Anoos, M.A. (2024). Eects of
humic and fulvic acids on forage production and grain quality of triticale
under various soil salinity levels. Cereal Research Communications.
https://doi.org/10.1007/s42976-024-00609-0
Muscolo, A., Francioso, O., Tugnoli, V., & Nardi, S. (2007). The auxin-like
activity of humic substance is related to membrane interactions in carrot
cell cultures. Journal of Chemical Ecology, 33(1), 115-129. https://doi.
org/10.1007/s10886-006-9206-9
Nardi, S., Pizzeghello, D., Muscolo, A., & Vianello, A. (2002). Physiological eects
of humic substances on higher plants. Soil Biology and Biochemistry,
34(11), 1527-1536. https://doi.org/10.1016/S0038-0717(02)00174-8
Pizzeghello, D., Nicolini, G., & Nardi, S. (2001). Hormone-like activity of humic
substances in Fagus sylvatica forests. New Phyrologits,151(3), 647-657.
https://doi.org/10.1046/j.0028-646x.2001.00223.x
Rodríguez-Roque, M.K., Rojas-Grau, M.A., Elez-Martínez, P., & Martín-Belloso,
O. (2013). Soymilk phenolic compounds, isoavones, and antioxidant
activity as aected by in vitro gastrointestinal digestion. Food Chemistry,
136(1), 206-212. https://doi.org/10.1016/j.foodchem.2012.07.115
Rouphael, Y., Colla, G., & De Pascale, S. (2021). Sprouts, microgreens and edible
owers as novel functional foods. Agronomy, 11(12), 2568. https://doi.
org/10.3390/agronomy11122568
Sano, T., Kutsuna, N., Becker, D., Hedrich, R., & Hasezawa, S. (2009). Outward‐
rectifying K+ channel activities regulate cell elongation and cell division
of tobacco BY‐2 cells. The Plant Journal, 57(1), 55-64. https://doi.
org/10.1111/j.1365-313X.2008.03672.x
Seth, T., Mishra, G.P., Chattopadhyay, A., Roy, P.D., Devi, M., Sahu, A., &
Nair, R.M. (2025). Microgreens: Functional food for nutrition and
dietary diversication. Plants, 14(4), 526. https://doi.org/10.3390/
plants14040526
Sharma, S., Shree, B., Sharma, D., Kumar, S., Kumar, V., Sharma, R., & Saini, R.
(2022). Vegetable microgreens: The gleam of next generation superfoods,
their genetic enhancement, health benets, and processing approaches.
Food Research International, 155, 111038. https://doi.org/10.1016/j.
foodres.2022.111038
Tallei, T.E., Kepel, B.J., Wungouw, H.I.S., Nurkolis, F., & Fatimawali, A.A.A.
(2024). A Comprehensive review on the antioxidant activities and
health benets of microgreens: current insights and future perspectives.
International Journal of Food Science and Technology, 59(1), 58-71.
https://doi.org/10.1111/ijfs.16805
Verlinden, S. (2020). Microgreens: Denitions, product types, and production
practices, In I. Warrington (Ed.), Horticultural Reviews, 47 (pp. 85-124).
Wiley Online Library. https://doi.org/10.1002/9781119625407.ch3
Toscano, S., Cavallaro, V., Ferrante, A., Romano, D., & Patané, C. (2021). Eects
of dierent light spectra on nal biomass production and nutritional
quality of two microgreens. Plants, 10(8), 1584. https://doi.org/10.3390/
plants10081584
Wang, K., Li, Y., Zhang, Y., Lou, X., & Sun, J. (2022). Improving myobrillar
proteins solubility and thermostability in low-ionic strength solution:
A review. Meat Science, 189, 108822. https://doi.org/10.1016/j.
meatsci.2022.108822
Xiao, Z., Lester, G.E., Luo, Y., & Wang, Q. (2012). Assessment of vitamin and
carotenoid concentrations of emerging food products: edible microgreens.
Journal of Agricultural and Food Chemistry, 60(31),7644-7651. https://
doi.org/10.1021/jf2057816
Xiao, Z., Raush, S.T., Luo, Y., Sun, J., Yu, L., Wang, Q., Chen, P., Yu, L., &
Stommel J.R. (2019). Microgreens of Brassicaceae: Genetic diversity
of phytochemical concentrations and antioxidant capacity. LWT –
Food Science and Technology, 101, 731-737. https://doi.org/10.1016/j.
lwt.2018.10.076
Zhang, P., Zhang, H., Wu, G., Chen, X., Gruda, N., Li, X., & Duan, Z. (2021).
Dose-dependent application of straw-derived fulvic acid on yield and
quality of tomato plants grown in a greenhouse. Frontiers in Plant
Science, 12, 736613. https://doi.org/10.3389/fpls.2021.736613