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Evaluation of moringa (Moringa oleifera Lam.) leaf extract on bioactive compounds of lettuce (Lactuca sativa L.) grown under glasshouse environment
⁎Corresponding author at: University of Debrecen, Faculty of the Agricultural and Food Sciences and Environmental Management, Horticulture Department, H-4032 Debrecen, room 69, Böszörményi Street 138, Hungary. arshad.yaseen@agr.unideb.hu (Arshad Abdulkhalq Yaseen)
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Received: ,
Accepted: ,
This article was originally published by Elsevier and was migrated to Scientific Scholar after the change of Publisher.
Peer review under responsibility of King Saud University.
Abstract
This study aimed to assess the novelty of moringa (Moringa oleifera Lam.) leaf extract (MLE) as a natural biostimulant on bioactive compounds in lettuce. In autumn season of 2019 and 2020, MLE (6%) was foliar sprayed onto lettuce grown under glasshouse conditions. Vitamin C, total polyphenols, total chlorophyll content and nitrate accumulation in three lettuce cultivars (May King, Kobak, and Great Lakes) and some correlations among the bioactive compounds were assessed. The results in 2019 show that there was no significant differences in the total polyphenols (mg GAE 100 g−1 FW) and vitamin C (mg 100 g−1 FW) between treated and untreated plants, while in 2020 significantly higher vitamin C (mg 100 g−1 FW) and total polyphenols (mg GAE 100 g−1 FW) were recorded in the plants treated with 6% MLE. In contrast, in 2019 application of 6% MLE lowered nitrate content (mg kg−1) by 13% in May King, 23% in Kobak, and 44% in Great Lakes. Similarly, in the season of 2020 foliar application of 6% MLE reduced the leaf nitrate content by 16% in May King, 19% in Kobak, and 44% in Great Lakes. In the mean of varieties, the only significant difference in total polyphenol and vitamin C was in the Great Lakes variety grown in 2020 whereas no differences were found for the experiment of 2019. In the mean of years, the only statistical difference was recorded for the nitrate content in control of the Great Lakes variety by 44% lower nitrate in the treated plants with 6%MLE. Positive correlation was found between vitamin C and total polyphenol content, whereas nitrate had a negative regression slop to vitamin C and total polyphenol content in both experiments. The major total chlorophyll differences were found in the experiment of 2019, although no significant differences were found for the SPAD value of the experiment of 2020. In our 2 years study, there was a positive correlation between vitamin C and total polyphenol while negative correlation was found between vitamin C and total polyphenol to nitrate content in all the lettuce cultivars.
Keywords
Lettuce quality
Bioactive compounds
Lettuce cultivars
Moringa leaf extract
Nitrate accumulation
1 Introduction
Lettuce is the worlds’ most popular consumed fresh leafy vegetable. It plays a critical role in the human diet and benefits for weight losses due to its low calory and high dietary fiber content (10 kcal [60 kJ]/100 g, FW) (Mampholo et al., 2016), appreciable useful compounds like phenols, vitamin A, C and E, carotenoids, iron and calcium (Romani et al., 2002; Szeto et al., 2002). Polyphenols in lettuce leaves contain many antioxidant properties which provide better human health (Lima et al., 2014; Wang et al., 1998). Many studies have described the health benefits of lettuce consumption to humans and rats in the risk reduction of cardiovascular diseases (Nicolle et al., 2004; Serafini et al., 2002).
However, leafy vegetables are considered to accumulate higher nitrate than other vegetables, e.g. bulbs, tubers, and roots (Maynard et al., 1976; Santamaria, 2006), which may threaten human health while converting to nitrite and nitric oxide by salivary enzymes and oral bacteria (Alexander et al., 2008; Khan et al., 2015). So that, the Food and Agriculture Organization (FAO) and the European Commission (EC) have provided the daily acceptance of nitrate intake of 0–3.7 mg kg−1 body weight (Santamaria, 2006). Similarly, the U.S. Environmental Protection Agency (EPA) has set up an acceptable daily consumption of 7.0 mg kg−1 body weight (Mensinga et al., 2003).
Vegetables have a short lifecycle and need a higher amount of fertilizer during the growing season in comparison to cereals and fruit trees, thus they need more fertilizer and intensive care till they reach the harvest. Lactuca sativa is a soil nutrient-depleted vegetable since it grows fast, produces a very shallow root system. So that, it requires adequate levels of fertilizer during its grown season (Thorup-Kristensen, 2001). Although, excessive fertilization of the lettuce field may cause many issues to the environment as water quality throughout leaching and runoff, eutrophication, greenhouse effect, and acid rain (Gastal and Lemaire, 2002; Heckman, 2007; Heckman et al., 2003; Wang et al., 2002) and harmful effects on human health (Ikemoto et al., 2002; Liu et al., 2014).
Fertilizers and pesticides cannot play the entire role in this concomitant challenge that the agriculture sector is facing in the future of the fast global population and environmental impacts on the ecosystem. In this sense, biostimulants are the main focussed product by scientists as an appropriate alternative for the vegetable growers besides the fertilizer and pesticides to extra care of their plants. Biostimulants are mostly made from plant extracts which are used for multipurpose reasons in crop production to alleviate plant abiotic stress (Colla and Rouphael, 2015), nutrient use efficiency, and growth improvement (Rouphael and Colla, 2020).
The reduction of nitrate accumulation in our daily food is extremely important because the nutritional product quality is accounted as the global food security where it is strategically important for the governments all around the world (Bian et al., 2020). High nitrate content or nitrate accumulation in leafy vegetables especially lettuce is a major issue facing quality assistance of the vegetable producers. Thus, plant biostimulants are important not only as a safe material to the environment and human health, but reducing fertilizer inputs to the crops with the same or even better yield and quality (Bulgari et al., 2015).
Moringa leaf extract (MLE) is a recently concerned plant extract that is applied to the plants as an environmentally friendly and safe biostimulant since it contains many beneficial bioactive compounds, macro and micronutrients, minerals, plant hormones vital amino acids, and vitamins (Rady et al., 2013; Sohaimy et al., 2015; Yaseen and Takácsné-Hájos, 2020). The benefits of MLE as a growth enhancer, improve yield and quality has been found on many different crops, green bean (Elzaawely et al., 2017), wheat (Ju et al., 2019), organic fennel (Abdel-Rahman and Abdel-Kader, 2020; El-Serafy and El-Sheshtawy, 2020), freesia plant (Ahmad et al., 2019), mandarin (Nasir et al., 2016), pepper (Aluko, 2016). MLE is also considered as an alternative to inorganic fertilizer (Saini et al., 2016), and effective biostimulant to reduce plant stress (drought and salinity) of many crops (Lucini et al., 2015; Zulfiqar et al., 2020).
However, very little research is available on the influence of moringa leaf extract in improving the quality and nitrate reduction of different lettuce cultivars. Thus, these researches aimed to evaluate the role of moringa leaf extract (MLE) in improving some bioactive components in three lettuce verities grown under glasshouse conditions.
2 Materials and methods
2.1 Growth conditions and plant materials
In September 2019 and 2020, seeds of May King, Kobak, and Great Lakes lettuce varieties were sown under the optimal weather condition with 70 ± 5 % relative humidity, 25/15 ± 2 ℃ day/ night temperature under glasshouse of research field at the University of Debrecen, Hungary. After about 35 days of the seed sowing, the seedlings were transplanted to the controlled environment under glasshouse weather conditions with 10/14-h day/night photoperiod, 50–75% RH, 20/13 day/night temperature (Figs. 1 and 2). The seedlings were transplanted in the prepared glasshouse soil which the soil composition is shown in Table 1. In our experiment, 40 seedlings of the same development were transplanted per variety. They were divided into two groups (treated and untreated groups). Two weeks after transplanting, the seedlings were foliar sprayed with 6% MLE every two weeks intervals, where the control plants sprayed with distilled water only. Harvesting began 45 days after transplanting. The whole heads of the lettuce plants were carefully taken out for the sample assessment. The outer, middle and inner leaves of each plant were mixed and directly transferred to the lab for total polyphenol (mg GAE 100 g−1 FW), vitamin C (mg 100 g−1 product), and nitrate content (mg kg−1) measurements.
The average temperature and humidity during the lettuce growing season of 2019.
The average temperature and humidity during the lettuce growing season of 2020.
Parameters
Value
pH value (KCl)
7.4
EC (mS cm−1)
0.11
Carbonated lime g kg−1
30.3
Organic carbon – humus content (%)
1.7
Phosphorus pentoxide (mg kg−1) (AL)
236.0
Potassium oxide (mg kg−1)
177.0
Nitrate and nitrite -N (mg kg−1) (KCl)
70.6
Sodium (mg kg−1) (AL)
42.0
Magnesium (mg kg−1) (AL)
239
Sulphur (mg kg−1) (KCl)
50.4
Manganese (mg kg−1) (EDTA)
22.0
Zinc (mg kg−1) (EDTA)
1.9
Copper (mg kg−1) (EDTA)
1.4
2.2 Moringa leaf extract (MLE) preparation
In August 2019 and 2020, young leaves were harvested from the moringa trees grown under a glasshouse at the agrar campus, University of Debrecen, Hungary. Moringa leaf extract was prepared from the fresh young leaves (about 40 days old). First, the leaves were washed and placed in a refrigerator at 4 °C for 24 h as described by Yaseen and Hájos (2020). Then the stored leaves were grinded with the amount of (1 L water/10 kg fresh material) using a kitchen blender then the mixture was squeezed and passed through muslin cloth based on the method by Foidl et al., (2001). Then the extract was placed in 8, 000 X g centrifuge for 15 min and the needed concentration of 6% MLE was prepared for the foliar spray treatment. Some mineral elements were measured (Table 2). The extract was then kept in the refrigerator at 4 ℃ till the plants ready to be foliar sprayed (Fig. 3).
MLE
Values (mg)
Phosphorus (P)
100
Potassium (K)
350
Sulphur (S)
267
Calcium (Ca)
326
Magnesium (Mg)
86.8
Sodium (Na)
11.4
Iron (Fe)
2.24
Cupper (Cu)
0.319
Zinc (Zn)
0.477
Boron (B)
0.581
Moringa leaf extract (MLE) preparation from fresh leaves Created with BioRender.com.
2.3 Determination of total polyphenol content
The edible lettuce leaves of the inner, middle, and outer leaves of the plant samples were mixed and taken to the lab immediately after harvesting to measure the bioactive compound assessments. Total polyphenols in mg GAE 100 g−1 fresh product were performed from the sample solutions using Folin–Ciocalteu reagent. Samples of the solution (5.0 mL) were diluted in distilled water. Only 2.0 mL of each diluted sample was mixed with 10 mL of distilled water and 1.0 mL of prepared Folin–Ciocalteu reagent and placed in 250 mL flask. The volumetric flask was filled with sodium carbonate solution Na2CO3 (Sigma-Aldrich Kft., Budapest, Hungary) at 29% (w/v). The absorbance was measured using a spectrophotometer at 720 nm. The results were given in gallic acid equivalent (GAE) of 50 mM–2.5 mM gallic acid [in 95% (vol/vol) methanol] based on the method by Ainsworth and Gillespie (2007).
2.4 Ascorbic acid (Vitamin C) measurement
Vitamin C content was determined by redox titration using iodine solution following the method described by Ciancaglini et al. (2001). ascorbic acid + I2 → 2 I− + dehydroascorbic acid
2.5 Nitrate determination
The nitrate content in lettuce leaves was measured based on the method by Kmecl et al. (2007) using Segmented Flow Analyzer (AA II, Bran + Luebbe) at the wavelength of 540 nm after the reduction in copper coated cadmium column (NO3– + 2e– ® NO2–) to form a diazo compound.
2.6 Statistical analysis
Data were subjected to one-way analysis of variance (ANOVA) at the confidence level P ≤ 0.05 using SPSS version 25 (SPSS Inc., Chicago, IL, USA). The means of 12 samples/replication were subjected to the independent sample t-test to determine significant differences between the two groups.
SPAD-502 (Spectrum Technologies) device was used to perform the correlation between chlorophyll content, nitrate and bioactive compounds in lettuce leaves based on the method by Madeira et al. (2003). The data were statistically analysed using pearson correlation method by Weinberg and Abramowitz (2008) at p ≤ 0.05 and p ≤ 0.01.
2.7 Fertilizer application
Unlike other vegetables, lettuce growth, yield and some chemical contents as nitrate content are most affected by nitrogen fertilizer. Based on the recommended nitrogen fertilizer for lettuce as discussed by Liu et al., (2014), the inorganic fertilizer FERTICARE 24–8-16 (NPK) was applied with the amount of 200 kg N ha−1 to our plants.
3 Results and discussions
3.1 Total polyphenol content (mg GAE 100 g−1 FW)
The influence of the application of moringa leaf extract varied considering the environmental factors and plant genotypes. In the autumn season of 2019, statistically higher total polyphenol was measured in untreated Great Lakes variety, while non of the other varieties (May King and Kobak) were significantly different. However, in the season of 2020, foliar spray with 6% MLE could significantly enhance the total polyphenol content in both May King and Great Lakes, whereas no significant difference was observed for the Kobak variety (Table 3). Except for genetic factors many other factors can change the polyphenol quantity in the plant leaves as leaf position (inner or outer leaves), agronomical factors as well as plant tissue type (green, yellow or white) (Bilyk and Sapers, 1985; Romani et al., 2002). Means in the same column followed by the same symbol (*) are not significantly different at probability level (p ≤ 0.05) according to the independent sample t-test
Lettuce cultivars
Total polyphenol content (mg GAE 100 g−1 FW)
Growing seasons
In the mean of years
Mean ± Std. Deviation
2019
Mean ± Std. Deviation2020
Mean ± Std. Deviation
Control
6% MLE
Control
6% MLE
Control
MLE
May King
37.65 ± 2.04
35.59 ± 1.53
51.63 ± 2.00
65.83 ± 10.00
44.64± 7.86
50.71± 17.75
Kobak
37.95 ± 1.00*
35.61 ± 1.00
65.40 ± 4.07
70.88 ± 2.00
51.67± 15.26
53.24± 19.36
Great Lakes
41.73 ± 1.01*
37.26 ± 0.97
55.69 ± 3.00
73.03 ± 5.00*
48.71± 7.90
55.14± 19.85
In the mean of varieties
39.11 ± 2.33
36.15 ± 1.33
58.93 ± 9.24
69.91*± 6.5
–
In general, plants in 2020 had more temperature fluctuation stress than in 2019, for example, a dramatic drop in temperature of roughly 5 °C occurred towards the end of November 2020 and continued until the end of the season (Figs. 1 and 2). Thus, this might be the reason for having higher total polyphenol in the experiment of 2020. Many research studies have been provided on the abiotic stress factor affecting total polyphenol in plants (Abdallah et al., 2013; Helyes et al., 2015). The temperature in 2020 was much fluctuated and decreased in the last two growing months (November and December) than in 2019, thus the polyphenol was also greater in 2020. This result agrees with the result by Boo et al., (2011) where they found higher total polyphenol and anthocyanin contents in lettuce grown under day/night temperature of 13/10 °C and 20/13 °C, followed by 25/20 °C and 30/25 °C.
Nasir et al., (2020) have detected the improvement of polyphenol in mandarin fruit by 1.4-fold in the trees treated with 3% MLE in comparison to untreated trees. This might be owing to the zeatin content of the moringa leaves (Basra et al., 2011) because this enhances the antioxidant content of the plants through the enzymes in the plant cells (Zhang and Ervin, 2004). Yasmeen et al., (2013) have also found a maximum value of total phenolic content in the wheat grown under moderate saline growth conditions.
Elzaawely et al. (2017) further mentions that the quality enhancement in plants treated with MLE is related to the hormon concentration in moringa leaves, particularly gibberellins (GA7).
In the mean of years, the total polyphenol content in lettuce leaves was significantly improved in the foliar sprayed plants by 8% compared to the control plants. Where in the mean of plant varieties, there were no significant differences in the season of 2019 among the varieties, while in 2020 plants foliar sprayed with 6% MLE were significantly affected by the biostimulants in comparison to non-treated plants (Table 3).
3.2 Vitamin C (mg 100 g−1 fresh weight)
The ascorbic acid content of the lettuce cultivars ranged from 4.81 to 10.3 mg100 g−1 FW in both seasons 2019 and 2020. The highest amount of the vitamin C (10.3 mg100 g−1) was found in the Great Lakes variety in 2020 in the plants foliar sprayed with 6% MLE, while the lowest was in the Kobak and May King treated with 6% MLE in 2019 (4.81 and 4.91 mg100 g−1) fresh product, respectively (Table 4). The experiment results indicate the variation of the vitamin C content under the influence of the interaction of plant variety and the treatments from seasons 2019 and 2020. Lower vitamin C was found in the treated plants with 6% MLE for the season of 2019, however statistically no significant difference was found between treated and untreated cultivars. This shows that the used plant biostimulant (6%MLE) could perfectly work on the plants in 2020 where the treated plants could improve better under less abiotic stress by producing higher vitamin C than in 2019. Plant biostimulants can improve growth and reduce abiotic stress in plants (du Jardin, 2015; Rouphael and Colla, 2018). Whereas, except for the Kobak cultivar there was a significantly higher vitamin C in May King and Great Lakes cultivars treated with 6% MLE in the grown season of 2020. Nasir et al., (2020) demonstrated the improvement of vitamin C by 2.3-fold in mandarin fruit treated with 3% MLE comparing to the control. Moringa leaf extract (MLE; 30 times diluted) resulted in the maximum ascorbic acid content in the wheat grown under highly saline (12 dS m−1) growth conditions (Yasmeen et al., 2013). In the mean of years, plants were not affected by 6% MLE for the vitamin C content. Similarly, in 2019 no significant results appear between treated and non-treated plants among the varieties. In contrast, in the mean of varieties in 2020 plants treated with 6% MLE recorded 26% greater vitamin C content than non-treated plants (Table 4). Means in the same column followed by the same symbol (*) are not significantly different at probability level (p ≤ 0.05) according to the independent sample t-test
Lettuce cultivars
Vitamin C (mg 100 g−1 fresh wieght)
Growing seasons
In the mean of years
Mean ± Std. Deviation
2019
Mean ± Std. Deviation2020
Mean ± Std. Deviation
Control
6% MLE
Control
6% MLE
Control
MLE
May King
5.85 ± 0.90
4.81 ± 0.96
6.03 ± 0.76
9.21 ± 0.09*
5.94 ± 0.75
7.01 ± 2.48
Kobak
5.83 ± 0.17
4.91 ± 0.94
8.87 ± 0.75
9.92 ± 1.01
7.35 ± 1.73
7.41 ± 2.87
Great Lakes
8.61 ± 1.17
8.30 ± 0.69
6.95 ± 0.15
10.30 ± 0.30*
7.78 ± 1.17
9.30 ± 1.19
In the mean of varieties
6.76 ± 1.57
6.01 ± 1.88
7.28 ± 1.36
9.81 ± 0.71*
–
3.3 Nitrate content in lettuce leaves (mg kg−1)
Nitrate content in lettuce leaves was varied from year to year and among the varieties. Many factors can influence these differences as genetic, environmental impacts as light, temperature, and CO2 differences between the seasons (Bian et al., 2020; Maynard et al., 1976; Prugar et al., 1991), as well as plant age – e.g. younger plants, contain higher nitrate content than older ones (Geiger et al., 1998) as well as agronomical factors (Al-Gburi, 2021). Our result indicates that factors (genetic and biostimulants) could mitigate the nitrate concentration in lettuce fresh mass. Nitrate accumulation was varied among different lettuce genotypes, lower in May King followed by Kobak and Great Lakes variety. This is similar to the result by Nádasy and Nádasy (2006) where they found that Great Lakes was more accumulating nitrate than Kobak and May King varieties in their pot experiment of adding 100 mg N-fertilizer per kg of soil.
In recent years, many researchers have reported the positive influence of plant biostimulants, organic extracts, amino acids, and vitamins on nitrate reduction in plants. Examples are given on reduction of nitrate in lettuce, swiss chard, spinach, and radish using protein hydrolysates as biostimulants (Colla and Rouphael, 2015). Also, Amanda et al., (2009) have shown the positive influence of plant spraying with the biostimulant of Actiwave®, Valagro SpA at a concentration of 6 mL m−2 on nitrate reduction of lettuce grown under controlled environment. Table 5 shows that in the season of 2019, foliar application of 6% MLE lowered the leaf nitrate content by 13% in May King, 23% in Kobak, and 44% in Great Lakes. Similarly, in the season of 2020 foliar application of 6% MLE reduced the leaf nitrate content by 16% in May King, 24% in Kobak, and 44% in Great Lakes. Other researchers have shown that biostimulants as double irrigation with 1.0% Bio-algeen S-90 (200 mL per plant) extracted from the brown seaweed and 0.3% Megagreen (100 mL m−2) reduced nitrate content in lettuce leaves (Dudaš et al., 2016). Our results agree with the result by Fu et al., (2017) where the increase of vitamin C content in lettuce leaves the nitrate level decreases. Based on our previous results of the experiment of (Yaseen and Takácsné-Hájos, 2021), the high amount of amino acid content in the MLE could also inhibit the nitric ion in the soil while being absorbed by the plant leaves. Means in the same column followed by the same symbol (*) are not significantly different at probability level (p ≤ 0.05) according to the independent sample t-test
Lettuce cultivars
Nitrate accumulation in lettuce leaves (mg kg−1)
Growing seasons
In the mean of years
Mean ± Std. Deviation
2019
Mean ± Std. Deviation2020
Mean ± Std. Deviation
Control
6% MLE
Control
6% MLE
Control
MLE
May King
784.00 ± 22.00*
692.00 ± 16.00
439.00 ± 3.00*
379.00 ± 15.00
611.50±
189.48535.50±
171.99
Kobak
950.00 ± 42.00*
772.00 ± 20.00
642.00 ± 3.00*
517.00 ± 8.00
796.00±
170.78644.50±
140.33
Great Lakes
1180.66 ± 13.50*
820.00 ± 22.00
702.00 ± 8.00*
489.00 ± 4.00
941.33±
262.36*654.50±
181.84
In the mean of varieties
971.55±
174.27*761.33 ± 58.48
594.33 ± 119.44*
461.66 ± 63.77
–
In the mean of varieties, the application of 6% MLE could significantly reduce nitrate content by 27.5% in 2019 and 27.8 % in 2020. Whereas in the mean of years, the only significant difference was in Great Lakes variety with the highest reduction (44%) which was followed by Kobak and May King with 24 and 14 % (Table 5).
3.4 Chlorophyll content in related to MLE treatment
Many studies have shown that moringa leaf extract (MLE) can improve photosynthetic pigments under normal and stress environmental conditions. Based on our two-year research, 6% MLE colud considerably enhance total chlorophyll content in several lettuce cultivars. In the season of 2019, all the cultivars produced significantly higher chlorophyll content in the treated plants compared to untreated plants, whereas in the 2020 experiment, the treated lettuce cultivars produced higher chlorophyll pigment content, but there was no significant difference between control and treated plants with 6% MLE (Table 6.). The pigment concentration was varied among the lettuce varieties. For example, the Great Lakes variety had the greatest pigment concentration among the lettuce cultivars, which is owing to the fact that this variety is inherently greener in color, whilst others are lighter green to yellow. Similar findings were made by Ahmad et al. (2019), whom discovered a substantial increase in total chlorophyll in freesia (bulbous cut flower) plants. Other researchers show that foliar application of MLE dramatically increase phenolic antioxidants, other biochemicals, and total chlorophyll content in spinach (Spinacia oleracea L.) leaves (Aslam et al., 2016). Also, Elzaawely et al. (2017) discovered a substantial increase in chlrophyll a, b, and carotinoides in snap beans treated with 1:20 and 1:30 foliar sprays of MLE across two growth seasons. Similarly, our findings show that plants treated with 6% MLE had considerably greater chlorophyll pigmentation than untreated plants over the course of years. Means in the same column followed by the same symbol (*) are not significantly different at probability level (p ≤ 0.05) according to the independent sample t-test
Treatments
Cultivars
Total chlorophyll
(SPAD value)
Growing Seasons
Mean of Years
2019
2020
untreated
May King
13.16
19.43
16.30
Kobak
16.23
20.30
18.26
Great Lakes
32.76
36.06
34.41
treated
May King
17.56*
23.33
20.45*
Kobak
23.90*
20.56
22.23*
Great Lakes
40.03*
38.10
39.06*
3.5 Correlation & regression analysis results
Our findings reveal that the primary bioactive compounds in the 2019 trial showed a positive correlation with SPAD values. Total polyphenol, for example, showed a strong positive connection with vitamin C and Nitrate, but no relationships with SPAD value (chlorophyll content) were identified. Vitamin C, on the other hand, exhibited a positive association with nitrate content and SPAD value of 0.757** at p ≤ 0.05 for 2019 and 2020. Some of the experiment's 2020 results were entirely different from those in 2019. For example, there was no connection between vitamin C, nitrate, and SPAD (Table 7). In the experiment of 2019, vitamin C had a positive correlation with nitrate content at p ≤ 0.01, whereas in the season of 2020 there was no or negative correlation between vitamin C and nitrate content at (-) 0.172. The strongest correlation among bioactive chemicals was discovered during the growth season of 2019, but minimal correlation was observed during the growing season of 2020, this might be due to meteorological variations, particularly temperature and light. **Correlation is significant at the 0.01 level (1-tailed). *Correlation is significant at the 0.05 level (1-tailed).
Years
Total polyphenol
Vitamin C
Nitrate
SPAD
Total polyphenol
2019
1.000
2020
1.000
Vitamin C
2019
0.469*
1.000
2020
0.757**
1.000
Nitrate
2019
0.562*
0.488*
1.000
2020
−0.082
−0.172
1.000
SPAD
2019
0.279
0.708**
0.170
1.000
2020
0.096
0.143
0.309
1.000
In the mean of years, total polyphenol and vitamin C had a negative correlation regression with the nitrate content in lettuce cultivars at R2 = 0.532 and R2 = 0.234 respectively. Nonetheless a positive correlation regression equation between vitamin C and total polyphenol was found for the mean of the experiment of 2019 and 2020 at R2 = 0.6207 (Fig. 4).
Regression equation results of the studied data for the mean of bioactive compound value of 2019 and 2020.
4 Conclusion
In the consideration of climatic and genetic variables, Moringa (Moringa oleifera Lam.) leaf extract (MLE) can be a reliable source of safe and natural biostimulants to improve certain bioactive components and reduce nitrate in the edible part of lettuce. Our experiment gives a safe and simple method for farmers to obtain the extract, in which they may grow the tree in their field and make the extract for their plants. The use of plant biostimulants (6% MLE) decreased nitrate accumulation in the lettuce cultivars grown in autumn 2019 and 2020. By reducing nitrate concentration in plant leaves, the leaf extract can play a significant role in lowering human health risks as nitrite intake via vegetable consumption. Also, Whereas vitamin C and total polyphenols showed very little improvement. This extract is critical for reducing nitrate without losing other quality factors such as bioactive components (vitamin C and polyphenols).
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
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