Comprehensive Assessment of Brassica juncea variety rugosa (Pahari Rai) Accessions from the Sub-Himalayan Region: Phytochemical, Antioxidant, Enzymatic, Mineral, and Fatty Acid Profiling

doi:10.21203/rs.3.rs-4340286/v1

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Comprehensive Assessment of Brassica juncea variety rugosa (Pahari Rai) Accessions from the Sub-Himalayan Region: Phytochemical, Antioxidant, Enzymatic, Mineral, and Fatty Acid Profiling

https://doi.org/10.21203/rs.3.rs-4340286/v1

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Brassica juncea var. rugosa (Pahari rai), a broad leafy cruciferous vegetable is an essential and easy source of nutrition with numerous health-promoting factors. The phytochemical and antioxidant activity of leaf extracts showed the highest phenolic, flavonoid, ODP, and tannin content observed in the EEC-25 leaf methanolic extract. The highest protein content in EEC-25 followed by IC-524259 and IC-350800. The foremost concentration of carbohydrates was observed in germplasms IC-363758 (31.47±0.31 mg glucose g^-1), IC-410471 (31.12±0.18 mg glucose g^-1) and EEC-25 (26.45±0.19 mg glucose g^-1). In-vitro antioxidant potential viz., total antioxidant content was observed highest in EEC-25 methanolic leaf extract (41.91±0.28 mg AAE g^-1), FRAP activity in EEC-25 (5.91±0.68 mg AAE g^-1), maximum DPPH free radical scavenging activity in IC-597933 (IC₅₀=21.45±3.4 μg mL^-1), ABTS free radical scavenging activity in IC-524259 (94.17±0.48 μg mL^-1), superoxide radical scavenging activity of methanolic extract of PR-15 (IC₅₀=124.37 μg mL^-1), highest metal ion chelating activity of EEC-25 methanolic extract (IC₅₀=52.22 μg mL^-1), maximum reducing power activity in EEC-25 and IC-399839, all being significantly different (p≤0.05) from each other. Positive correlations have been observed among the total antioxidant, total phenolic, total flavonoid content, ODP, DPPH, and FRAP. Micronutrient analysis Pahari rai leaf was observed to be a rich source of essential minerals such as Mn, Fe, Zn, Ca, Cu, and Mg. Pahari rai was observed to be a rich source of MUFA and PUFA. An essential metric for assessing the nutritional value of various edible oils is the ω6:ω3 ratio was observed to be maximum in IC-350800 (2.53), IC-338535 (2.34), and IC-410471 (2.19). The multiutility of this leafy vegetable against numerous pathological ailments was attributed to the pharmacological activities of phytochemicals and to the development of functional food for human consumption.

Biological sciences/Biochemistry

Biological sciences/Biological techniques

Brassica juncea variety rugosa

Phytochemicals

Antioxidant

Correlation

mineral

Leafy vegetables are a cost-effective source of nutrient that everyone can adopt. Brassica juncea var. rugosa, popularly known as broad leafy mustard belongs to the family Cruciferae, also known as Brassicaceae. It is a tall, slow-growing annual green leafy vegetable (Rabi season: October-March) with large leaves whose color ranges from light green to deep purple. It is native to Central and Eastern Asia. This leafy mustard is a staple crop, grown and supplied mostly in Asian nations such as India, China, Japan, Nepal, and Bhutan. In India, it is mainly cultivated in in small patches in the home backyard, agricultural land and mountainous regions including Uttarakhand, Himachal Pradesh, Arunachal Pradesh, Nagaland, Meghalaya, Mizoram, Assam and Manipur. Popularity of this leafy vegetable owes to its ease of cultivation, low risk of production, traditionality and versatility of applications.

The crop is grown by direct sowing or transplantation (Parajuli 2015). The leaves of well grown rugosa are 40–50 cm long and 20–25 cm wide having flat petioles, smooth to crumpled margins, fleshy and non-hairy (Rauniyar and Bhattarai 2017; Adhikari et al. 2022). Enriched with water content, fiber, vitamins along with several phytochemicals and nutrients, rugosa leaf is widely considered a wholesome food (Pant et al. 2020). Being one of the mustard varieties, Brassica juncea var. rugosa is rich in nutrient, such as polyphenols and glucosinolates that holds value in promoting human health and are commonly eaten in the form of pickle (Harbaum et al. 2008; Lin et al. 2011).

Phytochemicals also known as secondary metabolites are biologically active molecules synthesized by plants through primary or secondary metabolism that aid in the defensive mechanisms of the plant against infection. Recent research suggests that consuming plants high in polyphenols can significantly reduce the development of cardiovascular disease, neurological illness, cancer and diabetes (Chauhan et al. 2012). Brassica juncea var. rugosa is surplus in many such phytochemical compounds that exhibit several medicinal and biological properties, along with high protein and carbohydrate content (Moirangthem and Praveen 2020). It includes considerable levels of iron, calcium, protein, as well as Vitamins A, B, C and E (Pant et al. 2020). Due to their great nutritional value and disease-fighting capabilities, green mustard leaves can be a nutritious substitute for the majority of leafy winter vegetables.

It is a promising low-cost source of critical nutrients, especially beneficial for vulnerable groups such as women and developing children. In response to this potential, germplasms from diverse hilly regions in various Indian states were meticulously collected. Subsequently, a comprehensive analysis involving phytochemical, antioxidant, mineral, and fatty acid characterization was conducted to pinpoint. The ultimate goal is to leverage these findings to enhance people's nutritional status and overall well-being. Pahari rai's significance lies in its role as an edible mustard species that has been consistently exploited for its nutraceutical attributes, holding immense potential for future direct applications in healthcare products. The detailed insights into the nutraceutical properties derived from this research have the potential to revolutionize the development of health-enhancing products, providing accessible and cost-effective solutions to address nutritional deficiencies and improve public health.

If not neutralized, free radicals can damage different cells and organs in both humans and plants, increasing their vulnerability to both biotic and abiotic stresses, leading to various degenerative illnesses. An innate or built-in antioxidant system can scavenge these free radicals contributing directly towards plant defense system and promote human health as well by restraining the degenerative ailments such as cardio-vascular diseases, cancer and even reducing the effects of aging.

Plant Material and Extraction

The details of the plant material that have been used for secondary metabolite profiling, enzymatic activity, mineral analysis, and fatty acid profiling of Brassica juncea var. rugosa is provided in Table 1. For the preparation of methanolic extracts of leaf, 1 g of dried leaf powder was dissolved in 100 mL 70% methanol and kept in an orbital shaker at 37°C for 24 hours. The solution was filtered using Whatman filter paper no. 1 and then the filtrate concentrated in rotavapor at 55°C (Vaijanathappa et al. 2008). The collected concentrate was re-suspended in absolute methanol to the final concentration of 1 g mL^− 1.

Table 1

*Brassica juncea* var. rugosa germplasm from various sub-Himalayan regions of India
S.No.	Accessions	Cultivar Name	District	State	Latitude	Longitude
1	IC-338751	Peli sarson	Champawat	Uttarakhand	29° 20' 9.9528'' N	80° 5' 27.6936'' E
2	IC-524259	Rai	Pauri	Uttarakhand	30° 09' 10.30" N	78° 46' 37.56" E
3	IC-338535	Lai	Almora	Uttarakhand	29° 35' 39.0804'' N	79° 39' 14.0148'' E
4	EEC-25	Lahi	Ranikhet	Uttarakhand	29° 38' 60.00" N	79° 25' 12.00" E
5	IC-399826	'Chhanakti' sarson	Chamba	Himachal Pradesh	32° 33' 19.12" N	76° 07' 35.29" E
6	IC-399839	Desi sarson	Kangra	Himachal Pradesh	32° 5' 59.2944'' N	76° 16' 8.7744'' E
7	IC-399880	Desi sarson	Kullu	Himachal Pradesh	31° 57' 28.2636'' N	77° 6' 34.0524'' E
8	IC-363758	Kodra	Sirmaur	Himachal Pradesh	30° 33' 46.2456"	77° 28' 12.7092" E
9	IC-597917	Tusut	East Siang	Arunachal Pradesh	28° 4′ 12″ N	95° 19′ 48″ E
10	IC-597873	Ogiyi	West Siang	Arunachal Pradesh	28° 23' 59.99" N	94° 32' 59.99" E
11	IC-417128	Pettu (Lai patta)	Upper Siang	Arunachal Pradesh	28° 36′ 37.33″ N	95° 2′ 51.11″ E
12	IC-597933	Sarson	Lower Subansiri	Arunachal Pradesh	27° 47' 59.99" N	93° 35' 59.99" E
13	IC-298019	Antum	Serchhip	Mizoram	23° 20' 29.9688'' N	92° 51' 0.8172'' E
14	IC-410471	Tryso	Ri-Bhoi	Meghalaya	25° 53' 14.9676" N	91° 52' 15.0996" E
15	IC-350800	Janem	Jaintia Hill	Meghalaya	25° 27′ 0″ N	92° 12′ 0″ E
16	IC-276011	Rayo	Tinsukia	Assam	27° 29′ 22.07″ N	95° 21′ 36.52″ E
17	IC-413486	Kala sarisha	Dhalai	Tripura	23° 54′ 58.82″ N	91° 53′ 16.15″ E
18	IC-597821		Jhansi	Uttar Pradesh	25° 30′ 0″ N	78° 30′ 0″ E
19	Pusa Saag		Pantanagr	Uttarakhand	28° 58′ 12″ N	79° 24′ 36″ E
20	PR-15 (Kranti)		Pantnagar	Uttarakhand	28° 58′ 12″ N	79° 24′ 36″ E

Phytochemical estimation

Total phenolic content

The total phenolic content was determined by Folin-Ciocalteau reagent (FCR) based method (McDonald et al. 2001). Briefly, 0.5 mL of methanolic extracts were dried in water bath. Subsequently 1 mL of DW was added followed by 0.5 mL of 50% (v/v) FCR and 1 mL Na₂CO₃. The tubes were then incubated for 1 hr in dark. Gallic acid was used as standard and concentration was measured by UV-Vis spectrophotometer at 725 nm.

Total flavonoid content

The total flavonoid content was determined by the method of Mandal et al. (2009). 200 µL leaf extract was diluted with methanol upto 1 mL followed by 4 mL of DW. Thereafter, 0.3 mL each of 5% Sodium nitrite and 10% AlCl₃ were added and the tubes were left at RT for 5 minutes. AlCl₃ forms stable complexes with C-4 keto groups and C-3 or C-5 hydroxyl groups of flavones and flavonols. The reaction was terminated by the adding 2mL of 1M NaOH in each tube and absorbance of colored solution was recorded at 510 nm. The concentration of total flavonoids was calculated as mg quercetin equivalent g^− 1 extract (mg QE g^− 1), from the standard curve of quercetin.

Orthodihydroxy phenol

The content of the orthodihydroxy phenol (ODP) was estimated by Arnow’s reagent (Arnow 1937). Briefly, 0.2 mL extract was added to 1 mL 0.05N HCl. Sequentially add 1 mL of Arnow’s reagent, 10 mL DW and 2 mL of 10 N NaOH. The tubes were incubated for 2 minutes and absorbance was taken at 515. The concentration of ODP was calculated from the standard curve of catechol.

Total tannin content

Tannin quantification was done by Folin-Denis method given by Bae et al. (1993). 0.2 mL extract was added to 0.5 mL Folin-Denis reagent followed by 1 mL saturated Na₂CO₃. The volume of the reaction mixture was made up of 10 mL using DW and incubated for 30 min at RT. Tannins reduce phosphotungstomolybdic acid to a blue color solution under alkaline condition. The intensity of the blue color was read at 760 nm. The amount of tannin was estimated as mg tannic acid equivalent g^− 1 extract (mg TAE g^− 1) from the calibration curve of the tannic acid.

Total Soluble Protein estimation

The quantitative analysis of the protein content in fresh leaves was determined according to Lowry et al. (1951). FCR gives a blue-colored complex when react with protein which is absorbs at 660 nm. A standard curve was prepared using different concentrations of BSA.

Total carbohydrate content

Phenol-sulphuric acid method was performed to quantify the total carbohydrate content in fresh leaves of Brassica juncea var. rugosa (Hossain et al. 2015). The concentration was measured spectrophotometrically at 490 nm with glucose as standard.

In-vitro Antioxidant activity

Determination of Total antioxidant activity

The total antioxidant activity was evaluated by phosphomolybedinum method. The plant extract reduces Mo (VI) to green phosphate Mo (V) complex under acidic pH. (Prieto et al. 1999). 1 mL of freshly prepared phosphomolybdate reagent was added to 1 mL of leaf extract and incubated at 95º C for 90 min. Once the reaction mixture cooled to RT, the absorbance was recorded at 695 nm. The total antioxidant activity of the extract was expressed as mg ascorbate equivalents g^− 1 of extract.

Ferric reducing antioxidant power (FRAP)

The FRAP assay was determined spectroscopically at 593 nm by creating a complex with Fe(III)-TPTZ (Benzie and Strain 1996). This assay is based on the reduction of 2,3,5-triphenyl-1,3,4-triaza-2-azoniacyclopenta-1,4-diene chloride-ferric iron (Fe(III)-TPTZ) complex to the blue ferrous form (Fe (II)-TPTZ) at low pH. Briefly, FRAP reagent was pre-warmed at 37°C then 1.8 mL was added to 100 µL extract followed by 100 µL distilled water. The antioxidant potential of extracts was evaluated from the calibration curve of ascorbic acid prepared in methanol.

DPPH radical scavenging activity

The free radical scavenging capacity of leaf extracts was determined using DPPH according to Yen and Duh 1993 with some modification. DPPH free radical absorbs at 517 nm i.e., purple hue and addition of substrate causes decolorization of purple color. In short, 1mL of methanolic solution containing varying concentrations of leaf extracts or ascorbic acid was mixed with 0.5 mL of (0.5 mM) methanolic solution of DPPH and incubated for 30 min in dark at RT. The free radical scavenging activity was calculated by the equation:

$$DPPH Scavenging \left(\%\right)=\left[1-\left(\frac{{Sample}_{Abs}}{{Control}_{Abs}}\right)\right]\times 100$$

Superoxide anion radical scavenging assay

Superoxide anion radicals are one of the ROS resulting from various metabolic processes in vivo. The superoxide anion scavenging ability of methanolic leaf extracts was evaluated following the method of Nishikimi et al. (1972). The active compounds in plant extract quenches anionic radicals inhibiting reduction of nitro blue tetrazolium (NBT) to purple formazan. 3 mL of tris-HCl (16 mM, pH-8.0) was added to a premix containing 1 mL of 50 µM NBT and 1 mL of 78 µM NADH. The reaction was started by adding 1 mL of 10µM phenazine methosulfate. Thereafter, different concentrations of plant extract (100–500 µg mL^− 1) was added and the reaction mixture was incubated for 5 minutes at 25°C. The absorbance was read at 560 nm, decreasing absorbance correlates to increasing superoxide anion scavenging activity of extracts. The percentage inhibition of superoxide anion was calculated using given formula:

$$Superoxide radical Scavenging \left(\%\right)=\left[1-\left(\frac{{Sample}_{Abs}}{{Control}_{Abs}}\right)\right]\times 100$$

Metal ion Chelating activity

The metal ion chelation ability of the extract was estimated according to Decker and Welch (1990). Briefly, 1 mL of 0.1 mM FeSO₄ was mixed with 1 mL of extract at various concentrations. Subsequently 1 mL of 0.25 mM ferrozine was added followed by 10 min incubation at RT. Ferrozine chelates Fe (II) to form a pink-colored ferrozine-Fe (II) complex. Plant extract competes with ferrozine, restricting initial reaction thus resulting in a decrease in red color intensity. The absorbance was measured at 562 nm with EDTA as a reference and the percentage inhibition of ferrozine-Fe II complex formation was given by the following equation:

$$Ferrous ion Chelating Activity \left(\%\right)=\left[1-\left(\frac{{Sample}_{Abs}}{{Control}_{Abs}}\right)\right]\times 100$$

ABTS Free Radical Scavenging Activity

The free radical scavenging activity of plant samples was performed by ABTS radical cation decolorization assay (Re et al. 1999). 7 mM ABTS was mixed with 2.45 mM potassium persulfate (1:1) and allowed to react by incubating in the dark at RT for 12–16 hr which resulted in ABTS^·+ radical production. This solution was diluted with methanol till an absorbance of 0.700 ± 0.01 units at 734 nm was obtained. 5 µL of plant extract was added to 3.995 mL of diluted ABTS^·+ solution and allowed to react for 30 min. The absorbance was measured at 734 nm and percentage inhibition was calculated using the formula.

$$ABTS scavenging effect=\frac{AB-AA}{AB}*100$$

AB = absorbance of ABTS radical + methanol

AA = absorbance of ABTS radical + sample extract/standard

Enzymatic Analysis

Peroxidase determination

Peroxidase act as antoxidant enzyme. 100 mg of fresh leaf tissue was homogenized with 1.5 ml ice-cold 100 mM sodium phosphate buffer (pH 7.5) containing 1 mM EDTA, 1% PVP and 10 mM β-mercaptoethanol. The homogenate was centrifuged at 10,000 RPM for 15 min at 4°C and supernatant was collected for enzymatic assay. A reaction mixture was prepared containing 3 mL of 50 mM guaiacol in 0.1 M phosphate buffer (pH 6.5), to this 0.1 mL of enzyme extract was added. The reaction was initiated by adding 0.1 mL of 800 mM H₂O₂ and a change in absorbance was recorded at 470 nm for 3 min at an interval of 30 sec (Shannon et al. 1966).

Superoxide dismutase determination

Superoxide dismutase is also an antioxidant enzyme that catalyzes the dismutation of superoxide to H₂O₂ and O₂. The enzyme was extracted the same as peroxidase. For SOD quantification, 1.5 mL of 100 mM tris HCl buffer (pH 8.2), 0.5 mL of 6 mM EDTA, 1 ml of 6 mM pyrogallol solution, and 0.1 mL of enzyme extract were added into the cuvette. Change in absorbance was recorded at 420 nm at an interval of 30 seconds up to 3 min (Marklund and Marklund 1974).

Mineral Composition in Leaves

The mineral analysis of Mn, Zn, Pb, Fe, Mg, Cd, Cu, Ni, and Ca in the leaf was estimated through atomic absorption spectroscopy (AAS) after digestion of dried and powdered leaf samples (Moscow and Jothienkatachalam 2012). The elemental analysis was based on the absorbance of the species at its resonance wavelength.

Micronutrient (NPK) Analysis

For the analysis of nitrogen and potassium, dried leaf samples were ground and then digested by H₂SO₄ and H₂O₂ on an open digestion furnace at 380°C followed by extraction with 4 M HNO₃ for 4 h. The nitrogen content in the digested solution was determined by the Kjeldahl method according to Snell and Snell (1959) while potassium content was determined by flame photometric method (Bhargava and Raghupathi 1994). Phosphorus content was estimated by the molybdate-vanadate method. The orthophosphates obtained in triacid digestion react with ammonium molybdate, and ammonium vanadate in an HNO₃ medium producing a yellow color complex which absorbs at 470 nm. The concentration was estimated against a standard curve of phosphorous (Jackson 1973).

Fatty acid profiling in seeds of Brassica juncea var. rugosa germplasms

Fatty acid profile is an indicator of the quality and stability of the oil. Oil was extracted from seed by cold percolation method. 20 mg of oil sample were transferred into the soxhlet extraction flask. Toluene was mixed with dried methanolic H₂SO₄ in the ratio of 99:1. Nitrogen was passed for 2 min to get rid of moisture. The sample was refluxed for 12 hrs at 50°C and then cooled down. Further, 4% NaCl and 4% KCl solution and 20mL of n-Hexane were added to it. The n-hexane layer was vortexed and separated out. Further 4mL Na₂CO₃ (4%)/KCl (4%) was added and again vortexed and the n-hexane layer was separated out. 1-1.5 µL of the upper layer was subjected to gas chromatography. The injection volume was 1-1.5 µL and the carrier gas was nitrogen. The flow rate was 40–50 mL mL^− 1 with split injection mode. A flame ionization detector was used for the determination of the analyte. The temperature of the detector was kept between 250–300°C (Christie 1990).

Total Phenolic Content, total flavonoid, orthodihydroxyphenol (ODP), Total Tannin

Phenolic and flavonoids serve as key indicators of the antioxidant capacity of plant extracts, while ODP and tannin contribute to their characteristic astringency and medicinal properties. There was a significant difference (p < 0.05) in total phenolic, flavonoid, ODP, and tannin. Figure 1 shows the various concentrations of phenolic, flavonoid, ODP, and tannin in methanolic extracts. Out of 20 samples methanolic extracts of EEC-25 (6.45 mg/g), IC-524259 (5.45 mg/g), IC-338751 (4.89 mg/g), and IC-597933 (4.99 mg/g) showed the higher concentration of phenolic content whereas rest of the samples have almost similar concentration ranged from 2.41–3.40 mg/g. The maximum flavonoid concentration in EEC-25 and Pusa saag were 6.32 and 5.11 mg/g, respectively. EEC-25 (4.66 mg/g), IC-399839 (3.31 mg/g), and IC-597821 (3.08 mg/g) were found to be the highest amount of ODP. The foremost tannic content was observed in EEC-25 (3.46 mg/g), IC-338751 (2.22 mg/g), and IC-597917 (2.13 mg/g).

Phenolic compounds include a benzene ring and one or more hydroxyl groups. A study by Moirangthem and Praveen (2020) depicted that the total phenolic, flavonoid, and Tannin of methanolic extracts of B. juncea var. rugosa leaves were 6.52 mg/g, 1.25 mg/g, and 2.64 mg/g respectively. The range of the total phenol concentration was identified as 4.68–5.47 mg/g and the ODP concentration from 0.40–0.99 mg/g. Phenolic compound serve as both radical scavengers and metal chelators due to its perfect structural chemistry. Flavonoids varied from 3.10–4.88 mg/g (Pant et al. 2020). The total flavonoid content in Brassica juncea var. gemmifera was reported to be 4.02 mg/g (Sun et al. 2018). ODP is an essential component of plant polyphenols. In a study reported by Parihar et al. (2012) ODP content ranged from 0.36–0.48 mg/g. In a study by Mobeen et al. (2021), the concentration of tannins ranged from 0.3–0.5 mg/g in Brassica juncea leaves which is lower than in the present study. Tannins were in charge of astringency and served to defend plants from insects and pests by binding to the digestive proteins of insects to generate an insoluble complex that rendered the enzymes inactive (Rao et al. 2017).

Total Soluble Protein and Total Carbohydrate Content

The protein content of B. juncea var. rugosa is shown in Fig. 2. There was a significant difference (p < 0.05) in total soluble protein concentration expressed as µg BSA g^− 1 in leaves of different accessions. The protein content observed to be maximum in EEC-25 (400.97 ± 16.5 µg BSA g^− 1) followed by IC-524259 (380.09 ± 17.22 µg BSA g^− 1), and IC-350800 (379.23 ± 18.26 µg BSA g^− 1). But the lower concentration of protein was observed in genotypes IC-417128 (209.07 ± 10.73 µg BSA g^− 1), IC-597933 (231.37 ± 23.12 µg BSA g^− 1), and IC-298019 (237.95 ± 19.76 µg BSA g^− 1). Total soluble proteins in fresh samples of B. juncea var. rugosa were determined by Moirangthem and Praveen (2020) by Bradford technique and it was found to be 2.2 mg/g. The soluble protein content in the individual edible parts ranged from 38.81–91.10 mg/g in Brassica juncea var. gemminfera (Sun et al. 2018). According to a study, the total protein concentration in B. juncea leaves varied from 0.6–0.9 mg/g (Mobeen et al. 2021).

The total carbohydrate content of B. juncea var. rugosa is shown in Fig. 3. There was a significant difference (p < 0.05) in total carbohydrate concentration expressed as mg glucose g^-1 in leaves of different accessions. The foremost concentration of carbohydrates was observed in genotypes IC-363758 (31.47 ± 0.31 mg glucose g^-1), IC-410471 (31.12 ± 0.18 mg glucose g^-1), and EEC-25 (26.45 ± 0.19 mg glucose g^-1). Whereas, the lowest concentration of carbohydrates was observed in IC-399839 (5.06 ± 0.25 mg glucose g^-1), IC-338535 (7.63 ± 0.23 mg glucose g^-1), and IC-597821 (7.60 ± 0.10 mg glucose g^-1). The carbohydrate content of fresh leaf samples of B. juncea var. rugosa was determined to be 16.66 mg/g (Moirangthem and Praveen 2020). In a study by Mobeen et al. (2021) the carbohydrate content ranged from 0.47–0.89 mg/g in leaves of B. juncea. Green vegetables, when consumed alongside other carb-rich foods like cereals, are acknowledged as a notable source of soluble carbohydrates that the body primarily utilizes for energy provision, contributing to substantial energy levels facilitating metabolism and the assimilation of meals (Saha et al. 2015).

In-vitro Antioxidant Activity

Total antioxidant content

Total antioxidant content of B. juncea var. rugosa was expressed as mg ascorbic acid equivalent (AAE) g^− 1 extracts. Total antioxidant content of all the accessions differed significantly (p < 0.05) with highest being in EEC-25 (41.91 ± 0.28 mg AAE g^− 1) followed by IC-399839 (39.96 ± 0.28 mg AAE g^− 1), and IC-276011 (36.2 ± 0.45 mg AAE g^− 1) as shown in Fig. 4. A significant amount of antioxidant activity in green vegetables is an indicator of disease prevention. A similar study done by Pant et al. (2020) reported antioxidant content of Brassica juncea variety rugosa from the lowest at 14.94 mg/g to the highest at 20.09 mg/g.

Ferric ion reducing antioxidant power (FRAP)

Ferric reducing activity was expressed as mg AAE g^− 1 and it was observed that it varied significantly (p < 0.05) among all the accessions as detailed in Fig. 5. The highest FRAP activity (5.91 ± 0.68 mg AAE g^− 1) was found in the EEC-25 leaf extract followed by that of IC-597917 (2.36 ± 0.06 mg AAE g^− 1) extract. The FRAP activity reported in our study is higher than what is reported by Deng et al. (2013) in Brassica juncea that is 8.28 ± 0.58 µmol/g.

ABTS Free Radical Scavenging Activity

The free radical scavenging activity of all the methanolic extracts were expressed as percentage inhibition against ABTS. IC-524259 (94.17 ± 0.48%) showed the highest ability to quench ABTS, followed by IC-399880 (92.91 ± 0.62%), IC-597917 (93.83 ± 0.62%), IC-597873 (93.09 ± 0.77%), and Pusa saag (94.01 ± 0.62%) (Fig. 6). Khanam et al. (2012) reported the percentage inhibition of Brassica rapa subsp. Chinensis (Pak choi) (96.97%) which corresponded with the values reported in our study. In another study by Fernández León et al. (2014) the ABTS activity was found to be 56.62 mg/100g in the petroleum ether extract of Brassica oleracea L. convar. capitata var. aabauda. This may indicate that the free radical scavenging activity was better retained in methanolic extracts.

DPPH, Superoxide anion radical, and metal ion chelation scavenging activity

Table 2

IC₅₀ of different antioxidant activities in leaf extracts of *Brassica juncea* var. rugosa
Accessions	DPPH scavenging (µg mL^− 1)	Superoxide Anion Radical Scavenging (µg mL^− 1)	Metal ion chelation (µg mL^− 1)
IC-338751	(353.05 ± 4.07)^e	(24.38 ± 1.56)^s	(283.91 ± 5.16)^f
IC-524259	(837.08 ± 5.60)^a	(1241.35 ± 3.08)^d	(202.70 ± 5.17)^l
IC-338535	(112.84 ± 3.66)ⁿ	(523.05 ± 2.49)ⁿ	(205.97 ± 5.01)^kl
EEC-25	(676.05 ± 4.58)^b	(732.28 ± 2.53)ⁱ	(52.22 ± 4.48)^o
IC-399826	(503.89 ± 4.20)^d	(1404.19 ± 3.82)^b	(213.56 ± 3.49)^k
IC-399839	(77.76 ± 4.04)^o	(433 ± 3.52)^o	(365.34 ± 4.51)^b
IC-399880	(285.49 ± 3.52)^f	(1696.30 ± 4.48)^a	(222.28 ± 4.03)^j
IC-363758	(625.31 ± 4.05)^c	(382.98 ± 3.08)^p	(203.64 ± 3.54)^l
IC-597917	(124.52 ± 4.55)^l	(1092.52 ± 2.99)^f	(384.25 ± 4.53)^a
IC-597873	(51.98 ± 4.08)^p	(1024.35 ± 2.64)^g	(306.73 ± 5.09)^e
IC-417128	(72.85 ± 3.58)^o	(652.12 ± 2.02)^k	(336.92 ± 5.52)^c
IC-597933	(21.45 ± 3.48)^q	(1302.11 ± 3.79)^c	(184.93 ± 3.47)^m
IC-298019	(117.16 ± 5.25)^mn	(544.58 ± 2.53)^m	(284.04 ± 4.55)^f
IC-410471	(156.19 ± 4.01)^j	(326.40 ± 3.23)^q	(327.81 ± 5.55)^d
IC-350800	(178.70 ± 2.11)ⁱ	(15.91 ± 1.54)^t	(264.12 ± 2.49)^h
IC-276011	(148.18 ± 3.09)^k	(1131.93 ± 3.04)^e	(271.51 ± 3.63)^g
IC-413486	(235.06 ± 4.01)^g	(664.11 ± 2.99)^j	(243.05 ± 3.46)ⁱ
IC-597821	(123.53 ± 2.10)^lm	(590.48 ± 1.50)^l	(97.26 ± 4.10)ⁿ
Pusa Saag	(160.75 ± 2.50)^j	(961.95 ± 2.02)^h	(212.98 ± 6)^k
PR-15	(222.38 ± 2.56)^h	(124.37 ± 2.03)^r	(290.50 ± 4.03)^f

DPPH is a relatively stable organic radical that has been widely used to determine the antioxidant activity of single compounds, as well as different plant extracts. The DPPH free radical scavenging activity, superoxide anion radical scavenging, and metal ion chelating activity was expressed as IC₅₀ values of the methanolic extracts (Table 2). The IC₅₀ values for all the activity differed significantly (p < 0.05) among all the accessions. IC-597933 showed the maximum DPPH free radical scavenging activity with lowest IC₅₀ value (21.45 ± 3.4 µg mL^− 1) followed by IC-597873 (51.98 ± 4.08 µg mL^− 1) and IC-417128 (72.85 ± 3.58 µg mL^− 1). Whereas the superoxide radical scavenging activity was found to be maximum in IC-350800 with IC₅₀ value of 15.91 µg mL^− 1 followed by IC-338751 (24.38 ± 1.56 µg mL^− 1), lower than the reference standard i.e., ascorbic acid (41.46 µg mL^− 1). The IC₅₀ value for metal ion chelating activity was observed lowest in EEC-25 (52.22 µg mL^− 1) followed by that of IC-597821 (97.26 µg mL^− 1), lower than the standard EDTA (150.33 µg mL^− 1).

At a concentration of 400 µg mL^− 1, the DPPH scavenging percentages were 62.75, 59.61, 62.95, 67.68, 69.52, 64.49, 60.06, and 68.77 for methanolic extracts of IC-350800, IC-413486, IC-410471, IC-597873, IC-298019, Pusa saag, PR-15, and standard, respectively (Fig. 7). At a concentration of 200 µg mL^− 1 superoxide anion radical scavenging percentages were 58.33, 66.01, 64.75, and 60.52 for the methanolic extract of IC-338751, IC-410471, PR-15 and standard, respectively. At a concentration of 400 and 500 mg mL^− 1, the scavenging percentages of methanolic extracts of all the genotypes are nearly the same or less as compared to the standard (Fig. 8).

In the methanolic extract of all the genotypes, at all the concentrations from 100–500 µg mL^− 1, the metal ion chelating activity were lower and similar to the reference (Fig. 9).

In a similar study by Moirangthem and Praveen (2020) determined the DDPH activity in methanolic, ethyl acetate, petroleum ether, and chloroform extract of B. juncea var. rugosa. At 50 g mL^− 1 concentration the ethyl acetate, petroleum ether, and chloroform extract exhibited 41.14, 40.33, and 10.33% scavenging activity respectively. The leaf of B. juncea showed 95.55% DPPH free radical scavenging activity (Saha et al. 2015). The DPPH percentage inhibition of Brassica rapa subsp. Chinensis (Pak choi) were found to be (72.16 ± 8.35 mg g^− 1) (Khanam et al. 2012). Since phenolics are a significant component of overall antioxidant activity, a higher level of total phenols may be responsible for the excellent DPPH activity. No literature reports were available on the quantification of superoxide activity in B. juncea var. rugosa. But in similar studies by Jaiswal et al. (2011) among the different Brassica vegetable extracts, York cabbage showed the H₂O₂ scavenging capacity (1.15 ± 0.06 mg mL^− 1), followed by broccoli (1.29 ± 0.10 mg mL^− 1), Brussels sprouts (2.14 ± 0.02 mg mL^− 1) and white cabbage (2.84 ± 0.21 mg mL^− 1). In a study, various Brassica vegetables were studied for their ferrous ion chelating capacity. Among the vegetables studied, the Fe²⁺ chelating capacity was found in York cabbage (1.49 ± 0.03 mg mL^− 1), followed by broccoli (1.58 ± 0.10 mg mL^− 1), Brussels sprouts (2.14 ± 0.36 mg mL^− 1) and white cabbage (3.45 ± 0.37 mg mL^− 1). Plant extracts contain flavonoids, which are known to form complexed with metal ions and are hence liable for the ability to scavenge free radicals (Rice- Evans and Miller 1996).

Peroxidase and Superoxide Dismutase Enzymatic Analysis

Further investigations were performed to identify the levels of peroxidase and superoxide dismutase in crude extract. The highest peroxidase enzyme activity found in the IC-350800, IC-410471, IC-298019, IC-338535, were 0.268, 0.177, 0.163, 0.142 units min^-1 g^-1 Fresh Weight (FW), respectively. Whereas, the lowest enzymatic activity of IC-338751 (0.002 ± 0.0035 units min^-1 g^-1 FW), followed by IC-413486 (0.05 ± 0.005 units min^-1 g^-1 FW) was observed (Fig. 10). Peroxidase activity enhances the shelf life by inhibiting lipid peroxidation, yellowing and chlorophyll degradation, ethylene production, and senescence (Zhuang et al. 1995). However, our study does not attempt to identify the compounds responsible for observed antioxidant effects from B. juncea var. rugosa. In a study published by Singh et al. (2010) the peroxidase activity in Brassica oleracea var. capitata genotypes ranged from 415–5314, with a genotype mean of 1929 ± 410.97 µmol tetraguaiacol min^-1 g^-1 FW.

The enzymatic activity of superoxide dismutase was found to be highest in IC-338751, followed by IC-524259 (3.65 ± 0.04 units min^− 1 g^− 1 FW), and IC-338535 (3.35 ± 0.03 units min^− 1 g^− 1 FW). But the rest of the genotypes had a similar activity (Fig. 11).

Understanding the plant’s enzymatic antioxidant activity is important for a better plant defense system and longer shelf life. In a study by Singh et al. (2010) SOD activity in Brassica oleracea var. capitata lines, ranged from 4.86–7.63 with a mean value of 6.46 ± 0.25 unit of SOD min^− 1 g^− 1 FW. In a research by Gul et al. (2013), the levels of SOD in Brassica rapa L. crude extract, was found to be 1 mg mL^− 1, obtained in chloroform and ethyl acetate fraction from Brassica rapa L. (both at 190 U mL^− 1), while 5 mg mL^− 1 in aqueous fraction (215 U mL^− 1) and 10 mg mL^− 1 in aqueous and ethyl acetate fraction (both at 220 U mL^− 1).

Pearson-Correlation Analysis

The correlation analysis of the phytochemical, antioxidant and enzymatic analysis is depicted in Fig. 12. Total flavonoid content highly correlated with protein, FRAP, and ODP while moderately correlated with tannins whereas ODP positively correlated with total antioxidant, FRAP as well as tannins. Flavonoids reduce ferric ions, scavenge ROS and neutralize their harmful effects, thereby reducing oxidative stress, thus preventing cellular damage (Rice-Evans et al. 1997; Middleton et al. 2000). The strong positive correlation of total flavonoid content with FRAP attributes to the antioxidant activity of flavonoids. Since proteins are involved in key biological processes, the positive correlation between total flavonoid content and protein may indicate that flavonoids, being powerful antioxidants, help protect them from oxidative damage thereby maintaining their structural integrity and functionality (Amarowicz et al. 2008; Ebrahimzadeh et al. 2008). The structural similarities and antioxidant properties shared between flavonoids and ODPs explain the positive correlation between total flavonoid content and ODP levels.

Tannins, one of the major classes of phenolic compounds found in plants are known for their ability to bind and precipitate proteins, contributing to their astringent taste. Total phenolic content, commonly used as an indicator of the antioxidant capacity of plant extracts, is a measurement that encompasses various phenolic compounds, including tannins. Their positive correlation with ODP can be attributed to the fact that tannins contribute significantly to the overall phenolic content and exhibit strong antioxidant activity due to their ability to scavenge free radicals and chelate metal ions (El Gharras 2009).

Total phenolic content positively correlated with the DPPH and SOD assays as the phenolic compounds, such as flavonoids and tannins possess strong radical scavenging capabilities thus effectively neutralizing the DPPH radical. The ability of phenolic compounds to scavenge superoxide radicals and reduce oxidative stress, thereby supporting the activity of SOD (Martins et al. 2015). The protein content showed moderate correlation with DPPH assay. This positive correlation is because certain proteins like catalase and peroxidase, or peptides inherently exhibit antioxidant effects by catalyzing the breakdown of ROS, scavenging free radicals or chelating metal ions (Shahidi and Zhong 2008).

Positive correlation between total antioxidant activity and total phenolic content suggested that phenolics may be one of the major sources of these vegetables’ antioxidant potential. A significant positive correlation of total flavonoid content with ODP, tannins, FRAP, total antioxidant activity and protein infer that the selection of antioxidant rich lines should be based on total flavonoid content either alone or in combination with ODP, tannins, FRAP, total antioxidant activity and protein. Therefore, ODP, tannins, FRAP, total phenolic content and protein could serve as a reliable biochemical marker to identify the germplasms having higher activity of other antioxidants such as total antioxidant, DPPH and SOD.

Mineral composition in Brassica juncea var. rugosa leaves

Leafy mustard is a rich source of minerals, according to mineral profiling (Table 3). The concentration of copper was found to be highest in IC-338751 (0.069 ppm), EEC-25 (0.053 ppm), and IC-597873 (0.048). In IC-597917, PR-15, and IC-399880 the calcium concentration was found to be foremost 0.038, 0.034, and 0.026, respectively.

Manganese was found to be maximum in IC-413486 (0.284 ppm), PR-15 (0.278 ppm), and Pusa saag (0.267). IC-399880, IC-399826, and IC-597873 contain the fruitful amount of Zn which is 0.157, 0.134, and 0.135, respectively. In contrast, the Fe concentration in IC-524259 (5.321 ppm), IC-413486 (4.374 ppm), and IC-338751 (4.092 ppm) was found paramount. In IC-413486 (8.006 ppm), IC-399839 (7.292 ppm), and IC-597917 (6.889 ppm) the magnesium concentration was found to be maximum.

The detection of heavy metals such as Ni, Pb, and Cd (Table 2) in Brassica juncea var. rugosa leaves through AAS can be attributed to both environmental factors and the natural properties of the plant. Environmental contamination is a primary source of heavy metal accumulation in plants. Industries, vehicular emissions, agricultural practices, and improper waste disposal can release heavy metals into the environment, which are then absorbed by plants.

Table 3

Mineral Composition in leaves of *Brassica juncea* var. rugosa (ppm)
Accessions	Cu	Ca	Mn	Zn	Fe	Mg	Cd	Ni	Pb
IC-338751	0.069	0.009	0.202	0.118	4.092	5.042	0.014	0.033	0.248
IC-524259	0.044	0.016	0.199	0.133	5.312	4.561	0.005	0.068	0.453
IC-338535	0.038	0.015	0.188	0.122	3.951	4.886	0.001	0.081	0.109
EEC-25	0.053	0.015	0.198	0.14	4.02	5.37	0.006	0.108	0.512
IC-399826	0.032	0.013	0.196	0.134	3.898	5.321	0.01	0.04	0.181
IC-399839	0.035	0.017	0.195	0.14	3.611	7.292	0.017	0.09	0.023
IC-399880	0.022	0.026	0.188	0.157	3.249	4.674	0.021	0.062	0.016
IC-363758	0.03	0.03	0.195	0.115	3.226	4.202	0.059	0.088	0.482
IC-597917	0.034	0.038	0.21	0.111	3.655	6.889	0.064	0.15	0.128
IC-597873	0.048	0.002	0.224	0.135	3.53	6.559	0.074	0.141	0.34
IC-417128	0.031	0.007	0.207	0.109	3.372	5.512	0.08	0.11	0.1
IC-597933	0.01	0.009	0.206	0.068	3.703	4.339	0.086	0.132	0.112
IC-298019	0.033	0.012	0.246	0.124	3.363	5.628	0.085	0.133	0.216
IC-410471	0.016	0.018	0.243	0.077	3.45	4.013	0.103	0.165	0.158
IC-350800	0.028	0.018	0.246	0.119	3.096	5.158	0.111	0.083	0.148
IC-276011	0.023	0.022	0.248	0.114	3.251	4.983	0.105	0.092	0.227
IC-413486	0.021	0.023	0.284	0.112	4.374	8.006	0.12	0.084	0.151
IC-597821	0.023	0.026	0.273	0.095	3.4	5.608	0.113	0.131	0.116
Pusa Saag	0.017	0.026	0.267	0.09	3.13	5.676	0.113	0.121	0.183
PR-15	0.015	0.034	0.278	0.114	3.118	6.676	0.121	0.049	0.226

For women and growing children, iron is vital, and mustard green is the greatest solution to provide them with their daily needs. The B. juncea var. rugosa accessions possessed Fe ranging from 11.81–20.23 mg/100g.. Iron has significance in metabolic processes like DNA synthesis, respiration, and photosynthesis, as well as a crucial part in physiological processes and biochemical pathways, iron is an essential component of the enzyme cytochrome of the electron transport chain (Rout and Sahoo 2015). Zinc concentrations ranged from 13.6–2.73 mg/100g. Manganese levels in B. rugosa genotypes ranged from a high of 6.49 mg/100g to a low of 4.17 mg/100g (Pant et al. 2020). As per Saha et al. (2015) Fe, Zn, Cu, Mn, Ca, and Mg content in Brassica juncea were 118.50, 7.50, 2.60, 45.97, 36.6, 16.33 mg/100g, respectively. Optimum levels of potassium enhance iron absorption and are advantageous for those taking diuretics to control hypertension (Archana et al. 2013). Mustard green can supply a significant amount of calcium, which is similarly crucial for bone health, especially in women and growing babies. Calcium functions as a secondary messenger in numerous physiological and developmental processes, as well as its involvement in the formation of cell walls, stability of cell membranes, and how it responds to biotic stress by concentrating when necessary (Thor 2019). The human body requires copper for the production of enzymes, and it functions as biological electron carrier, whereas magnesium plays a pivotal role in calcium metabolism, regulating blood pressure and insulin release (Alinnor and Oze 2011).

In 2013, Babar et al. reported that zinc, a protein cofactor involved in the production of chlorophyll, growth regulation, and stem elongation, serves as a functional, structural, or regulatory cofactor of several enzymes. Magnesium plays a vital role in various biological processes, such as glycolysis, oxidative phosphorylation, nucleic acid synthesis, energy production, mitochondrial ATP synthesis, cell signaling, and activation of cAMP (Saris et al. 2000; Barbagallo and Dominguez 2007). Nickel is one of the micronutrients in plants that hydrolyze urea in plant tissues and it is part of the active site of the enzyme. The experimental results from the current study will be crucial for the creation of new Ayurvedic formulations and will be used to manage the dosage and determine the ratio of different active ingredients in each formulation.

Micronutrient (NPK) Analysis

The maximum concentration of Nitrogen (N) in EEC-25, IC-597917, and IC-410471 was 0.644%, 0.588%, and 0.504%, respectively. IC-524259 and IC-399839 showed the minimum amount of N. In the case of phosphorous (P) EEC-25 (0.751%), IC-298019 (0.772%), and IC-524259 (0.749%) had the paramount concentration. The lowest amount of P was found in IC-363758 (0.334%), and IC-399826 (0.315%). IC-413486 (0.099%), IC-597873 (0.097%), and IC-399839 (0.093%) had the maximum concentration of potassium (K). furthermore, PR-15 (0.047%), IC-597933 (0.055%), and IC-338535 (0.050%) showed the lowest concentration of K (Fig. 13).

A study by Pant et al. in 2020 determined the phosphorous content ranged from 513mg/100g to high as 720 mg/100g. Only one piece of literature available for the NPK content in B. juncea var. rugosa. But in a study by Devkota et al. (2020), the concentration of NPK was 1.08, 1.6, and 2.83 respectively. Nitrogen is a necessary component of plant elements such as proteins, amino acids, nucleotides, nucleic acids, and chlorophyll content, nitrogen is essential for crop development. Potassium regulates protein turnover, effect the Rubisco activity, nitrate reduction process, ribosome polypeptide syntheses, translational process, synthesis, and activity of nitrate red, affects glucose starch synthase, glucose pyrophosphorylase, and β-amylase (Berg et al. 2009). The deficiency of potassium in the plasma can cause gastrointestinal problems such as vomiting, diarrhea, increased renal loss, muscle weakness, ECG abnormalities, decreased reflex response, and cardiac arrhythmias in severe cases (Kardalas et al. 2018). Phosphorous is one of the crucial macronutrients for the plant, which is involved in nucleic acids, enzymes, coenzymes, nucleotides, phospholipids, cell organization, root development, stalk or stem growth and strength, root development, photosynthesis, respiration, macromolecular biosynthesis (Thakur et al. 2018).

Fatty acid, oil stability index, protein and glucosinolates profiling

The fatty acid composition, oil stability index (OSI), as well as the oil content, protein and glucosinolate (GSL) content in seeds of Brassica juncea var. rugosa accessions (Table 4). The saturated fatty acid palmitic acid (16:0) was highest in genotypes IC-338751 (3.4%) followed by EEC-25 (2.98%), IC-597821 (2.62%), and IC-413486. Stearic acid (18:0) was maximum in IC-298019 (0.94%), but IC-524259 (0.88%), and IC-597933 (0.88%) almost have the same amount of stearic acid but IC-597873 (0.11%), and PR-15 (0.12%) had the lowest amount was observed. In a study by Rai et al. (2018) on fatty acid composition of various Brassica genotypes in different cultivars of Brassica, Brassica juncea genotypes contained the highest levels of palmitic acid, as a saturated fatty acid. The study also reported that the total SFA content in various Brassica was less than 7%, which is regarded as appropriate for human consumption. Oleic acid one of the monounsaturated fatty acids was present foremost in IC-367658 (20.11%), IC-597917 (17.11%), IC-524259 (16.13%) whereas IC-276011 (9.9%), and PR-15 (9.3%) had the modest amount. IC-597873 (10.56%) and IC-417128 (10.65%) have almost the same amount of oleic acid observed. The oleic acid content in the Brassica genotypes varied from 0.80 to 48.7% in Brassica juncea, 16.15–37.98% in Brassica napus, and 6.21–16.15% in Brassica rapa, among other MUFAs (Rai et al. 2018). Since oleic acid improves the concentration of high-density lipoproteins and reduces the concentration of low-density lipoproteins in the blood, higher doses of oleic acid are thought to have nutritional value for human nutrition (Chang and Huang 1998). It was observed that eicosenoic acid was highest in IC-338535 (10.11%), IC-524259 (9.68%), and IC-597933 (9.58%). In a finding by Velasco et al. (1998), the eicosenoic acid concentration was 6.2% in B. napus. Erucic acid was paramount in PR-15 (59.55%), IC-597873 (58.47%), and IC-597821 (53.29%). The cardiac conductance in humans is hampered by higher erucic acid concentration in cooking oil, which raises blood cholesterol levels (Bozzini et al. 2007). However, numerous germplasm undertaken in the current study that have higher quantities of erucic acid will be crucial for a variety of industrial applications. Erucic acid-rich oil is utilized as a raw ingredient in the detergent, plastic, tannery, cosmetic, and polyester industries (Rakow and Raney 2003; Coonrod et al. 2008). Brassica breeding projects are currently concentrated on the establishment of zero erucic lines for nutritional purposes. In a study by Tripathi (2019), the erucic content in various Indian mustard genotypes ranged from 0.93–51.4%.

The utmost linoleic acid (omega-6) content was observed in EEC-25 (22.48%), IC-338751 (20.23%), and IC-399880 (20.22%). On the other hand, for linolenic acid (omega-3), among all twenty germplasm, IC-276011 (12.68%), IC-399826 (12.65%) and EEC-25 (12.18%) had highest amount. In a similar study in B. juncea, B. napus, and B. rapa genotypes, the respective linoleic acid content ranged from 11-45.3%, 18.57–26.93%, and 14.08–18.18% while, linolenic acid content ranged from, 11-26.72%, 9.99–17.23%, and 9.82–26.66% (Rai et al. 2018). In a study by El-Beltagi et al. (2010) the level of PUFAs in various cultivars of Brassica napus varied from 10.52–13.41 for linoleic acid and 8.83–10.32 for linolenic acid. Which is very similar to the current study.

The PUFAs are recognized as the long chain fatty acid with more than one double bond, that functions as precursors for the production of molecules with crucial physiological functions like prostaglandins, leukotrienes and thromboxane. PUFAs cannot be synthesized by the human body, hence they must be supplemented through diet. High concentrations of linoleic acid in edible oils are said to lower blood cholesterol and prevent atherosclerosis (Ghafoorunissa, 1994). Despite the health benefits of linolenic acid, it’s inclusion in the oil elevates the risk of rancidity and bad flavor (Sharafi et al., 2015). Therefore, its concentration in the cooking oil, should be low. According to this study, Brassica juncea var. rugosa germplasm with low erucic and high linoleic acid concentrations can be employed in Brassica breeding programs aimed at strengthening the grade of oil for dietary and industrial applications.

An essential metric for assessing the nutritional value of various edible oils is the ω6:ω3 ratio. The ω6:ω3 ratio was observed to be maximum in IC-350800 (2.53), IC-338535 (2.34), and IC-410471 (2.19). The SFA:PUFA ratio was found highest in both germplasms IC-338751 (3:34) and EEC-25 (3:34), followed by IC-399826 (3:32), IC-524259 (3:29). The ratio of SFA:MUFA:PUFA was observed highest in IC-597873 (1:47:14), IC-597917 (1:26:11), IC-363758 (1:23:11) while lowest in germplasms IC-410471 (1:17:8), IC-399826 (1:17:9) and EEC-25 (1:17:9). The oil stability index of IC-363758 (1.047), IC-597917 (0.940), and Pusa saag (0.896) were highest in these genotypes. In all the genotypes protein percentage ranged from 22–30. But the EEC-25 (123.96), IC-298019 (116.42) and IC-417128 (115.98) have the highest content of glucosinolates.

The lack of these essential fats in our diets may contribute to the expansion of numerous chronic diseases in humans, including heart failure, cancer, diabetes, asthma, depression, accelerated aging, Alzheimer’s, obesity, and arthritis (Simpoulos, 2002). Balanced diets high in MUFA and PUFA have been advised over saturated fats to lower plasma cholesterol levels. A similar finding in B. napus L. by El-Beltagi et al. (2010) depicted the ratio of ω6:ω3 ranged from 3.85–5.32. Which is very much similar to the several genotypes but in some genotypes, it was found to be much higher. The excessive concentration of PUFA in edible oils is more susceptible to oxidation, which can lower the quality of both the oil itself and the products made with it. Rancidity and the production of free radicals are mostly caused by the oxidation of the double and triple bond formation in PUFA. So, it means that those oil have maximum OSI, having a longer shelf-life. In a study by Chauhan et al. (2010) the OSI ranged from 0.40 (Kranti of Indian mustard) to 4.96 (GSC-5 of Brassica napus). The ω6:ω3 ratio ranged from 0.55–3.5, which is very low compared to the current study.

Table 4

Fatty acid, oil stability index (OSI), oil, protein, and glucosinolate (GSL) content in seeds of *Brassica juncea* var. rugosa
	SFA		MUFA			PUFA
Accessions	Palmitic	Stearic	Oleic	Ecosinic	Erucic	Linoleic (ω6)	Linolenic (ω3)	ω6:ω3	SFA:PUFA	SFA:MUFA:PUFA	OSI	Protein (%)	GSL
IC-338751	3.4	0.54	13.8	7.09	44.2	20.23	14.1	1.43	3:34	1:16:8	0.682	23.15	92.06
IC-524259	2.52	0.88	16.1	9.68	35.41	18.68	10.98	1.70	3:29	1:18:8	0.863	24.45	91.17
IC-338535	2.5	0.74	14.1	10.11	42.5	19.2	8.21	2.34	1:09	1:20:8	0.735	24.33	75.32
EEC-25	2.98	0.58	15.5	5.25	40.42	22.48	12.18	1.85	3:34	1:17:9	0.690	23.68	123.96
IC-399826	2.55	0.84	12.1	9.21	39.42	20.18	12.65	1.60	3:32	1:17:9	0.600	25.28	89.99
IC-399839	2.5	0.72	13.2	8.23	40.41	18.65	11.12	1.68	3:29	1:19:9	0.706	26.18	98.13
IC-399880	2.4	0.59	15.1	8.72	41.52	20.22	17.67	1.14	2:37	1:21:12	0.748	27.40	61.11
IC-363758	2.18	0.58	20.1	6.5	39.38	19.21	12.48	1.54	2:31	1:23:11	1.047	24.66	97.09
IC-597917	2.12	0.51	17.1	7.45	45.22	18.21	11.53	1.58	2:29	1:26:11	0.940	27.25	98.19
IC-597873	1.47	0.11	10.6	6.3	58.47	12.78	10.01	1.28	1:22	1:47:14	0.826	25.06	101.24
IC-417128	2.18	0.5	10.7	0.55	45.41	17.64	9.68	1.82	2:27	1:21:10	0.604	27.92	115.98
IC-597933	2.52	0.88	15.1	9.58	38.32	18.63	10.48	1.78	3:29	1:18:8	0.812	26.37	109.92
IC-298019	2.25	0.94	11.7	7.45	45.2	14.15	11.36	1.25	3:25	1:20:7	0.828	27.40	116.42
IC-410471	2.4	0.73	11.5	0.84	42.34	18.29	8.37	2.19	3:26	1:17:8	0.629	23.56	82.39
IC-350800	2.14	0.81	11.7	0.47	48.92	17.65	6.99	2.53	1:12	1:20:8	0.660	23.30	94.68
IC-276011	2.19	0.75	9.9	7.2	45.88	16.02	12.68	1.26	1:14	1:21:9	0.618	26.90	100.06
IC-413486	2.58	0.86	11.8	7.53	45.69	16.28	10.74	1.52	1:09	1:18:7	0.723	23.46	77.37
IC-597821	2.62	0.36	11.2	5.68	53.29	17.76	11.67	1.52	2:29	1:23:9	0.631	22.32	81.9
Pusa Saag	2.36	0.96	13.3	7.07	45.42	14.83	11.82	1.25	3:26	1:19:8	0.896	26.88	97.53
PR-15	2.13	0.12	9.3	5.21	59.55	15.22	6.94	2.19	1:11	1:32:9	0.611	22.72	102.24

Vegetables not only contribute essential carbohydrates, proteins and lipids but also serve as a rich source of antioxidant. Recent research highlights the significant role of Brassica in promoting health and mitigating the risk of chronic diseases and age-related disorders. The body’s metabolic processes generate free radicals, which are normally countered by both enzymatic and non-enzymatic antioxidants within the body’s innate defense system, as well as those obtained from fruits and vegetables in the diet. Vegetables, containing phytochemicals such as phenols, flavonoids, and tannins, play a crucial role in neutralizing free radicals. Remarkably, these phytochemicals exhibit strong free radical scavenging activities, offering protection against oxidative damage and reducing overall oxidative stress on cells. The present study focused on determining phytochemical content, in-vitro antioxidant activity, correlation analysis, mineral composition and fatty acid profiling of Brassica juncea var. rugosa. The results demonstrated that among the collected 20 genotypes, EEC-25 methanolic leaf extract exhibited the highest levels of total phenolic, flavonoid, orthodihydroxyphenol, and tannin content. Furthermore, it displayed remarkable antioxidant activity, determined by total antioxidant activity, FRAP activity, and reducing power, making it a potential candidate for functional food applications. The correlation analysis revealed interesting associations between phytochemicals, antioxidant activity, and enzymatic analyses, shedding light on the underlying mechanisms of antioxidant potential in these accessions. The fatty acid profiling of seeds provided valuable insights into their nutritional characteristics, with the ω6:ω3 ratio indicating IC-350800, IC-338535, and IC-410471 have better nutritional value for edible oil purposes. Furthermore, the oil stability index highlighted the suitability of IC-363758, IC-597917, and Pusa saag as potential sources of vegetable oil with good shelf-life. Overall, this comprehensive investigation contributes to a better understanding of the phytochemical and antioxidant potential of Brassica juncea var. rugosa (Pahari rai) accessions from the Indian sub-Himalayan region, providing valuable data for future breeding and crop improvement programs, and offering opportunities for developing functional foods and stable vegetable oils with potential health benefits.

Acknowledgment

The authors would like to extend sincere thanks to the National Bureau of Plant Genetic Resources, Delhi, and the Department of Genetics and Plant Breeding, Gobind Ballabh Pant University of Agriculture and Technology, Pantnagar for providing the seed material for the experimental trails. The first author is highly indebted to Dr. Usha Pant, Department of Genetics and Plant Breeding Gobind Ballabh Pant University of Agriculture and Technology, Pantnagar for their guidance.

AUTHOR INFORMATION

Authors and Affiliations

Department of Biochemistry, College of Basic Sciences and Humanities, G. B. Pant University of Agriculture and Technology, Pantnagar, Uttarakhand, 263145, India

Ankur Adhikari, Himanshu Punetha

CONTRIBUTIONS

Ankur Adhikari: data curation; data analysis, original draft

Himanshu Punetha: Review and editing

Corresponding author: Correspondence to Ankur Adhikari and Himanshu Punetha

CONFLICT OF INTEREST

The authors declare that there are no competing interests associated with this paper.

PERMISSION OR LICENSE

The permission and seed material were procured from the Germplasm Evaluation Division, ICAR-National Bureau of Plant Genetic Resources, New Delhi. A research paper was already been published in the Electronic Journal of Plant Breeding (Genetic diversity analysis using agro-morphological traits of Brassica juncea subspecies rugosa (Pahari rai) from North-Eastern Himalayan region)

STATEMENT OF EXPERIMENTAL RESEARCH

The collected seed material was cultivated at Norma E. Borlaug, Crop Research Centre of Govind Ballabh Pant University of Agriculture and Technology, Pantnagar, Uttarakhand. The study was in accordance with relevant institutional, national, and international guidelines and legislation.

DATA AVAILABILITY STATEMENT

All data generated or analyzed during this study are included in this published article

Adhikari A, Punetha H, Pant U (2022) Genetic diversity analysis using agro-morphological traits of Brassica juncea subspecies rugosa (Pahari rai) from North-Eastern Himalayan region. Electronic J of Plant Breeding 13(3):1-10. https://doi.org/10.37992/2022.1303.106.
Alinnor IJ, Oze R (2011) Chemical evaluation of the nutritive value of Pentaclethra macrophylla benth (African Oil Bean) Seeds. Pakistan J Ntr 10(4):355-359. https://doi.org/10.3923/pjn.2011.355.359.
Amarowicz R, Dykes GA, Pegg RB (2008) Antibacterial activity of tannin constituents from Phaseolus vulgaris, Fagoypyrum esculentum, Corylus avellana and Juglans nigra. Fitoterapia 79(3):217-219. https://doi.org/10.1016/j.fitote.2007.11.019.
Archana GN, Pradeesh S, Chinmayee MD, Mini I, Swapna TS (2012) Diplazium esculentum: a wild nutrient-rich leafy vegetable from Western Ghats. In Prospects in Bioscience: Addressing the issues. Springer, India. https://doi.org/10.1007/978-81-322-0810-5_35.
Arnow LE (1937) Colorimetric determination of the components of 3, 4-dihydroxyphenylalanine-tyrosine mixtures. J Biol Chem 118(2):531-537.
Babar H, Yusop MK, Saleem M (2013) Role of zinc in plant nutrition-a review. J Exp Agri Inter 3(2):374-391. https://doi.org/10.9734/AJEA/2013/2746
Bae HD, McAllister TA, Muir AD, Yanke LJ, Bassendowski KA, Cheng KJ (1993). Selection of a method of condensed tannin analysis for studies with rumen bacteria. J of Agri and Food Chem 41(8):1256-1260. https://doi.org/10.1021/jf00032a018.
Barbagallo M, Dominguez LJ (2007). Magnesium metabolism in type 2 diabetes mellitus, metabolic syndrome and insulin resistance. Archives of Biochem and Biophy 458(1):40-47. https://doi.org/10.1016/j.abb.2006.05.007
Benzie IF, Strain JJ (1996). The ferric reducing ability of plasma (FRAP) as a measure of “antioxidant power”: the FRAP assay. Analytical Biochem 239(1):70-76. https://doi.org/10.1006/abio.1996.0292
Berg WK, Cunningham SM, Brouder SM, Joern BC, Johnson KD, Volenec JJ (2009) Influence of phosphorus and potassium on alfalfa yield, taproot C and N pools, and transcript levels of key genes after defoliation. Crop Sci 49:974–982. https://doi.org/10.2135/cropsci2008.07.0395
Bhargava BS, Raghupathi HB (1994) Analysis of plant materials for macro and micronutrients. Methods of analysis of soils, plants, water and fertilizers 49-82.
Bozzini A, Calcagno F, Soare T (2007) “SINCRON”, A NEW Brassica carinata Cultivar For Biodiesel Production/“Sincron”, La Nueva Variedad De Brassica Carinata Para La Producción De Biodiesel/“Sincron” Un Nouveau Cultivar De Brassica Carinata Pour La Production De Biodiesel. Helia 30(46):207-214. https://doi.org/10.2298/hel0746207b
Chang NW, Huang PC (1998) Effects of the ratio of polyunsaturated and monounsaturated fatty acid to saturated fatty acid on rat plasma and liver lipid concentrations. Lipids 33(5):481-487. https://doi.org/10.1007/s11745-998-0231-9
Chauhan JS, Kumar S, Singh KH, Meena SS, Meena ML (2010) Oil and seed meal quality indices of Indian rapeseed-mustard varieties. J of Plant Biochem and Biotech 19(1):83-86. https://doi.org/10.1007/BF03323440
Chauhan PK, Jaryal M, Kumari K, Singh M (2012) Phytochemical and in vitro antioxidant potential of aqueous leaf extracts of Brassica juncea and Coriandrum sativum. Int J of Pharma Sci and Res 3(8):2862-2865.
Christie WW 1990 Preparation of Methyl Esters Part-1. Lipid Tech 2:48-49.
Coonrod D, Brick MA, Byrne PF, DeBonte L, Chen Z (2008) Inheritance of long chain fatty acid content in rapeseed (Brassica napus L.). Euphytica 164(2):583-592. https://doi.org/10.1007/s10681-008-9781-7
Decker EA, Welch B (1990) Role of ferritin as a lipid oxidation catalyst in muscle food. J of Agri and Food Chem 38(3):674-677. https://doi.org/10.1007/s10681-008-9781-7
Deng GF, Lin X, Xu XR, Gao LL, Xie JF, Li HB (2013) Antioxidant capacities and total phenolic contents of 56 vegetables. J. of Functional Foods 5(1):260-266. https://doi.org/10.1016/j.jff.2012.10.015.
Devkota C, Bhattarai BP, Mishra SR, Ghimire P, Chaudhari D (2020) Effect of integrated plant nutrient management on growth, yield and leaf nutrient status of broadleaf mustard (Brassica juncea var. rugosa). Horti Int J 4(3):78-81. https://doi.org/10.15406/hij.2020.04.00162
Ebrahimzadeh MA, Pourmorad F, Bekhradnia AR (2008). Iron chelating activity, phenol and flavonoid content of some medicinal plants from Iran. African J of Biotech, 7(18):3188-3192.
El Gharras H (2009) Polyphenols: food sources, properties and applications–a review. Int J of Food Sci and Tech, 44(12):251-2518. https://doi.org/10.1111/j.1365-2621.2009.02077.x
El-Beltagi HEDS, Mohamed AA (2010). Variations in fatty acid composition, glucosinolate profile and some phytochemical contents in selected oil seed rape (Brassica napus L.) cultivars. Grasas Y Aceites 61(2):143-150. https://doi.org/10.3989/gya.087009
Fernández León AM, Lozano RM, González GD, Ayuso YMC, Fernández León MF (2014) Bioactive compounds content and total antioxidant activity of two savoy cabbages. Czech J Food Sci 32(6):549-554.
Ghafoorunissa G (1994) Dietary fats/oils and heart diseases. Sustainability in oil seeds. Indian Society of Oil Seeds Res Hyderabad, 486-490.
Gul S, Ahmed S, Gul H, Shad KF, Zia-Ul-Haq M, Badiu D (2013) The antioxidant potential of Brassica rapa L. on glutathione peroxidase, superoxide dismutase enzymes and total antioxidant status. Romanian Rev of Lab Med 21:161-169. https://doi.org/10.2478/rrlm-2013-0008
Harbaum B, Hubbermann EM, Zhu Z, Schwarz K (2008) Impact of fermentation on phenolic compounds in leaves of pak choi (Brassica campestris L. ssp. Chinensis var. communis) and Chinese leaf mustard (Brassica juncea Coss). J of Agri and Food Chem 56(1):148-157. https://doi.org/10.1021/jf072428o
Hossain M, Ahmed K, Chowdhury F, Roksana K, Islam S, Barman A (2015) Experimental study on grain weight, moisture, ash, carbohydrates, protein, oil, total energy and minerals content of different varieties of rapeseed and mustard (Brassica spp.). Int J Sci Res Publ 5:394-400.
Jackson ML (1973) Soil chemical analysis. Prentice hall of India Pvt. Ltd., New Delhi, India 498: 151-154.
Jaiswal AK, Rajauria G, Abu-Ghannam N, Gupta S (2011) Phenolic composition, antioxidant capacity and antibacterial activity of selected Irish Brassica vegetables. Nat Prod Comm 6(9):1299-304. https://doi.org/10.1177/1934578X11006009
Kardalas E, Paschou SA, Anagnostis P, Muscogiuri G, Siasos G, Vryonidou A (2018) Hypokalemia: a clinical update. Endocr. Connect 7:135–146. https://doi.org/10.1530/EC-18-0109
Khanam UKS, Oba S, Yanase E, Murakami Y (2012) Phenolic acids, flavonoids and total antioxidant capacity of selected leafy vegetables. J of Fun Foods 4(4):979-987. https://doi.org/10.1016/j.jff.2012.07.006
Lin LZ, Sun J, Chen P, Harnly J (2011) UHPLC-PDA-ESI/HRMS/MS n analysis of anthocyanins, flavonol glycosides, and hydroxycinnamic acid derivatives in red mustard greens (Brassica juncea Coss variety). J of Agri and Food Chem 59(22):12059-12072. https://doi.org/10.1021/jf202556p
Lowry O, Rosebrough N, Farr AL, Randall R (1951) Protein measurement with the Folin phenol reagent. J of Bio Chem 193(1):265-275.
Mandal S, Hazra B, Sarkar R, Biswas S, Mandal N (2009) Hemidesmus indicus, an age old plant: study of its in vitro antioxidant and free radical scavenging potentials. Pharmacologyonline 1:604-617.
Marklund S, Marklund G (1974) Involvement of the superoxide anion radical in the autoxidation of pyrogallol and a convenient assay for superoxide dismutase. European J of Biochem 47(3):469-474.
Martins N, Barros L, Santos-Buelga C, Silva S, Henriques M, Ferreira IC (2015) Decoction, infusion and hydroalcoholic extract of cultivated thyme: Antioxidant and antibacterial activities, and phenolic characterisation. Food Chem 167:131-137. https://doi.org/10.1016/j.foodchem.2014.06.094
McDonald S, Prenzler PD, Antolovich M, Robards K (2001) Phenolic content and antioxidant activity of olive extracts. Food Chem 73(1):73-84. https://doi.org/10.1016/S0308-8146(00)00288-0
Middleton E, Kandaswami C, Theoharides TC (2000) The effects of plant flavonoids on mammalian cells: implications for inflammation, heart disease, and cancer. Pharmacological Rev 52(4):673-751.
Mobeen Wang X, Saleem MH, Parveen A, Mumtaz S, Hassan A, Yasin G (2021) Proximate composition and nutritive value of some leafy vegetables from Faisalabad, Pakistan. Sustainability 13(15):8444. https://doi.org/10.3390/su13158444
Moirangthem L, Praveen N (2020) Phytochemical analysis and antioxidant activity studies of leaf of Brassica juncea var. rugosa through sequential solvent extraction. Int J of Pharm Sci and Res 11(1):343-351
Moscow S, Jothivenkatachalam K (2012) Study on mineral content of some Ayurvedic Indian medicinal plants. Int J Pharma Sci and Res 3(2): 294-299.
Nishikimi M, Rao NA, Yagi K (1972) The occurrence of superoxide anion in the reaction of reduced phenazine methosulfate and molecular oxygen. Biochem and Biophy Res Comm 46(2):849-854. https://doi.org/10.1016/S0006-291X(72)80218-3
Pant U, Bhajan R, Singh A, Kulshesthra K, Singh AK, Punetha H (2020) Green leafy mustard: A healthy alternative. E J of Plant Breeding 11(01):267-270. https://doi.org/10.37992/2020.1101.045
Parajuli A (2015) Cultivation and Management practices of leafy vegetable in Nepal. Wordpress.com. https://parajulianish.wordpress.com/2015/02/13/cultivation-and-management-practices-of-leafy-vegetables-in-nepal/ Accessed 13 Feb 2015
Parihar PS, Prakash O, Punetha H (2012) Changes in metabolites of Brassica juncea (Indian mustard) during progressive infection of Alternaria brassicae. Nat Sci 10: 39-42.
Prieto P, Pineda M, Aguilar M (1999) Spectrophotometric quantitation of antioxidant capacity through the formation of a phosphomolybdenum complex: specific application to the determination of vitamin E. Anal Biochem 269(2):337-341. https://doi.org/10.1006/abio.1999.4019
Rai GK, Bagati S, Rai PK, Rai SK, Singh M (2018) Fatty acid profiling in rapeseed mustard (Brassica species). Int J Curr Microbiol App Sci 7(5):148-157. https://doi.org/10.20546/ijcmas.2018.705.019
Rakow G, Raney JP (2003) Present status and future perspectives of breeding for seed quality in Brassica oilseed crops. In Proc. 11^th Int. Rape Seed Congress, Copenhagen, Denmark (pp. 181-185).
Rao DE, Divya K, Prathyusha IVSN, Krishna CR, Chaitanya KV (2017) Insect-resistant plants. Current Dev in Biotech and Bioeng 47-74. https://doi.org/10.1016/B978-0-444-63661-4.00003-7
Rauniyar K, Bhattarai BP (2017) Growth, yield and soil nutrient status of broad leaf mustard (Brassica juncea var. rugosa) under integrated nutrient management. Nepalese J Agri Sci 15:98-106.
Re R, Pellegrini N, Proteggente A, Pannala A, Yang M, Rice-Evans C (1999) Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radical Bio and Med 26(9-10):1231-1237. https://doi.org/10.1016/S0891-5849(98)00315-3
Rice-Evans CA, Miller NJ (1996) Antioxidant activities of flavonoids as bioactive components of food. Biochemical Society Transactions, 24(3):790-795. https://doi.org/10.1042/bst0240790
Rice-Evans C, Miller N, Paganga G (1997) Antioxidant properties of phenolic compounds. Trends in plant Sci 2(4):152-159. https://doi.org/10.1016/S1360-1385(97)01018-2
Rout GR, Sahoo S (2015) Role of iron in plant growth and metabolism. Rev in Agri Sci 3:1-24. https://doi.org/10.7831/ras.3.1
Saha J, Biswal AK, Deka SC (2015) Chemical composition of some underutilized green leafy vegetables of Sonitpur district of Assam, India. Int Food Res J (Malaysia) 22(4):1466-1473
Saris NEL, Mervaala E, Karppanen H, Khawaja JA, Lewenstam A (2000) Magnesium: an update on physiological, clinical and analytical aspects. Clinica chimica acta 294(1-2):1-26. https://doi.org/10.1016/S0009-8981(99)00258-2
Shahidi F, Zhong Y (2008) Bioactive peptides. Journal of AOAC international 91(4):914-931. https://doi.org/10.1093/jaoac/91.4.914
Shannon LM, Kay E, Lew JY (1966) Peroxidase isozymes from horseradish roots: I. Isolation and physical properties. J of Bio Chem 241(9):2166-2172. https://doi.org/10.1016/S0021-9258(18)96680-9
Sharafi Y, Majidi MM, Goli SAH, Rashidi F (2015) Oil content and fatty acids composition in Brassica species. Inter J of Food Properties 18(10):2145-2154. https://doi.org/10.1080/10942912.2014.968284
Simopoulos AP (2002) The importance of the ratio of omega-6/omega-3 essential fatty acids. Biomed. & Pharmacotherapy 56(8):365-379. https://doi.org/10.1016/S0753-3322(02)00253-6
Singh BK, Sharma SR, Singh B (2010) Antioxidant enzymes in cabbage: variability and inheritance of superoxide dismutase, peroxidase and catalase. Scientia Horti 124(1):9-13. https://doi.org/10.1016/j.scienta.2009.12.011
Snell FD, Snell CT (1959) Colorimetric methods of analysis. D. van Nostrand. (Book)
Sun B, Tian YX, Jiang M, Yuan Q, Chen Q, Zhang Y, Tang HR (2018) Variation in the main health-promoting compounds and antioxidant activity of whole and individual edible parts of baby mustard (Brassica juncea var. gemmifera). RSC Adv 8(59):33845-33854. https://doi.org/10.1039/C8RA05504A
Thakur AK, Singh KH, Singh L, Nanjundan J, Khan YJ, Singh D (2018) SSR marker variations in Brassica species provide insight into the origin and evolution of Brassica amphidiploids. Hereditas 155(1):1-11. https://doi.org/10.1186/s41065-017-0041-5
Thor K (2019) Calcium—Nutrient, and messenger. Frontiers in Plant Sci 10:440. https://doi.org/10.3389/fpls.2019.00440 (Review Article)
Tripathi CSM (2019) Biochemical studies in Indian mustard (Brassica juncea L.) Czern and Coss for fatty acid profiling. Int J of Chem Sci 7(4):338-343.
Vaijanathappa J, Badami S, Bhojraj S (2008) In vitro antioxidant activity of Enicostemma axillare. J of Health Sci 54(5):524-528. https://doi.org/10.1248/jhs.54.524
Velasco L, Goffman FD, Becker HC (1998) Variability for the fatty acid composition of the seed oil in a germplasm collection of the genus Brassica. Genetic Res and Crop Ev 45(4):371-382. https://doi.org/10.1023/A:1008628624867
Yen GC, Duh PD (1993) Antioxidative properties of methanolic extracts from peanut hulls. J of the American Oil Chemists’ Society 70(4):383-386. https://doi.org/10.1007/BF02552711
Zhuang H, Hildebrand DF, Barth MM (1995) Senescence of broccoli buds is related to changes in lipid peroxidation. J of Agri and Food Chem 43(10):2585-2591. https://doi.org/10.1021/jf00058a006

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Comprehensive Assessment of Brassica juncea variety rugosa (Pahari Rai) Accessions from the Sub-Himalayan Region: Phytochemical, Antioxidant, Enzymatic, Mineral, and Fatty Acid Profiling

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Abstract

Figures

Introduction

Material and Methods

Plant Material and Extraction

Phytochemical estimation

Total phenolic content

Total flavonoid content

Orthodihydroxy phenol

Total tannin content

Total Soluble Protein estimation

Total carbohydrate content

Determination of Total antioxidant activity

Ferric reducing antioxidant power (FRAP)

DPPH radical scavenging activity

Superoxide anion radical scavenging assay

Metal ion Chelating activity

ABTS Free Radical Scavenging Activity

Enzymatic Analysis

Peroxidase determination

Superoxide dismutase determination

Mineral Composition in Leaves

Micronutrient (NPK) Analysis

Results and Discussion

Total Phenolic Content, total flavonoid, orthodihydroxyphenol (ODP), Total Tannin

Total Soluble Protein and Total Carbohydrate Content

In-vitro Antioxidant Activity

Total antioxidant content

Ferric ion reducing antioxidant power (FRAP)

ABTS Free Radical Scavenging Activity

DPPH, Superoxide anion radical, and metal ion chelation scavenging activity

Peroxidase and Superoxide Dismutase Enzymatic Analysis

Pearson-Correlation Analysis

Micronutrient (NPK) Analysis

Fatty acid, oil stability index, protein and glucosinolates profiling

Conclusion

Declarations

References

Additional Declarations

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