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ORIGINAL ARTICLE
Year : 2018  |  Volume : 14  |  Issue : 55  |  Page : 72-77  

A rapid high-performance liquid chromatography method for the simultaneous estimation of water-soluble vitamin in ten wild edible plants consumed by the tribal people of North-eastern Region in India


Department of Plant Chemistry, Botanical Survey of India, Acharya Jagadish Chandra Bose Indian Botanic Garden, Howrah, West Bengal, India

Date of Submission18-Oct-2017
Date of Acceptance20-Dec-2017
Date of Web Publication28-Jun-2018

Correspondence Address:
Tapan Seal
Department of Plant Chemistry, Botanical Survey of India, Acharya Jagadish Chandra Bose Indian Botanic Garden, Howrah - 711 103, West Bengal
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/pm.pm_481_17

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   Abstract 


Background: Bauhinia purpurea, Clerodendrum colebrookianum, Dillenia pentagyna, Diplazium esculentum, Houttuynia cordata, Oenanthe linearis, Potentilla lineata, Perilla ocimoides, Sonchus arvensis, and Zanthoxylum acanthopodium are potent wild edible plants and consumed by the tribal people of North-eastern region in India. Objective: A reversed-phase high-performance liquid chromatographic method using photodiode array detector with gradient elution has been developed and validated for the simultaneous quantitation of several water-soluble vitamins such as ascorbic acid (Vitamin C), thiamine (Vitamin B1), riboflavin (Vitamin B2), niacin (Vitamin B3), pantothenic acid (Vitamin B5), pyridoxine (Vitamin B6), and folic acid (Vitamin B9) in these ten wild edible plants. Materials and Methods: The chromatographic separation of vitamins was carried out on Acclaim C 18 column (5 μm particle size, 250 mm × 4.6 mm), Dionex Ultimate 3000 liquid chromatograph and detection was carried out at three different wave lengths (210, 245, and 254 nm) using a mobile phase of acetonitrile and aqueous trifluoroacetic acid (0.01% v/v) solution with gradient elution. Conclusion: The results of investigation showed that these plants are rich sources of vitamins, especially the B group of vitamins that can contribute immensely to nutrition, food security, and health and therapeutic benefits. The high percentage of recovery (98%–99%), low coefficient of variation (R2 > 0.99), low limit of detection, and limit of quantitation confirm the suitability of the method for simultaneous quantification of vitamins in these ten plants under investigation.
Abbreviations used: RSD: Relative standard deviation; LOD: Limit of detection; LOQ: limit of quantification; SEM: Standard error of mean; ND: Not detected; DPM: Dried plant material; TFA: Trifluoroacetic acid; v/v: Volume by volume; HPLC: High-performance liquid chromatography; nm: Nanometer; mg: Milligram; μg: Microgram; R2: Regression coefficient.

Keywords: B group vitamins, High-performance liquid chromatography analysis, Vitamin C, water-soluble vitamins, wild edible plants


How to cite this article:
Seal T, Chaudhuri K, Pillai B. A rapid high-performance liquid chromatography method for the simultaneous estimation of water-soluble vitamin in ten wild edible plants consumed by the tribal people of North-eastern Region in India. Phcog Mag 2018;14, Suppl S1:72-7

How to cite this URL:
Seal T, Chaudhuri K, Pillai B. A rapid high-performance liquid chromatography method for the simultaneous estimation of water-soluble vitamin in ten wild edible plants consumed by the tribal people of North-eastern Region in India. Phcog Mag [serial online] 2018 [cited 2019 Nov 22];14, Suppl S1:72-7. Available from: http://www.phcog.com/text.asp?2018/14/55/72/235281





Summary

  • A reversed-phase high-performance liquid chromatographic method using photodiode array detector with gradient elution has been developed and validated for the simultaneous quantitation of several water-soluble vitamins such as ascorbic acid (Vitamin C), thiamine (Vitamin B1), riboflavin (Vitamin B2), niacin (Vitamin B3), pantothenic acid (Vitamin B5), pyridoxine (Vitamin B6), and folic acid (Vitamin B9) in ten wild edible plants consumed by the tribal people of North-eastern region in India
  • The results of investigation showed that these plants are rich sources of vitamins, especially the B group of vitamins that can contribute immensely to nutrition, food security, and health and therapeutic benefits. The high percentage of recovery (98%–99%), low coefficient of variation (R2 > 0.99), low limit of detection, and limit of quantitation confirm the suitability of the method for simultaneous quantification of vitamins in these ten plants under investigation.



   Introduction Top


Vitamins are essential ingredients of food which are required in small amounts in the body on a regular basis to lead normal health and various physiological functions in the human body. They are widely distributed in natural food sources and can be easily introduced into the diets to satisfy daily needs. Vitamins are a group of organic compounds and can be categorized into two groups based on their solubility: fat-soluble vitamins and water-soluble vitamins. The former includes lipid-soluble Vitamins A, D, E, and K and other carotenoids, the latter is composed of water-soluble Vitamin C and eight B vitamins, namely thiamine (Vitamin B1), riboflavin (Vitamin B2), niacin (Vitamin B3), pyridoxine (Vitamin B6), pantothenic acid (Vitamin B5), biotin (Vitamin B7), folate (Vitamin B9), and cyanocobalamin (Vitamin B12).[1]

Measurement of vitamins in foods is complicated by many factors. It is very difficult to develop a single universal method for the simultaneous quantification of vitamin due to their diverse chemical structures and properties. Moreover, each vitamin can occur in different forms called vitamers that possess the same biological activity on ingestion. Vitamins often occur in food at relatively low levels and susceptible to degradation by exposure to light, air, heat, and high pH. Different methods including spectrophotometry, titration, high-performance liquid chromatography (HPLC), capillary electrophoresis (CE), high-performance thin layer chromatography (HPTLC) and microbiological assays have been used for the determination of water-soluble vitamins in various conditions. The most widely used methods for the determination of ascorbic acid together with B group vitamins are reversed-phase HPLC coupled with diode array detector, using a C18 column and aqueous-organic mobile phases, in acidic media.[2]

Plants rich in fruits, vegetables, whole grains, and legumes provide an abundance of vitamins and minerals to meet one's nutritional needs. The therapeutic potential of the vegetables is largely dependent on the presence of vital vitamins as well as micronutrients. Even though vitamin is required a small amount per day in health, it plays a vital role in our health. The consumption of leafy vegetables and fruits rich in vitamins is reported to reduce the risk of attack of various acute and chronic diseases.[3]

The wild plants have been a main source of food and medicine for tribal people. In most developing nations, numerous types of edible wild plants are exploited as sources of food to provide supplementary nutrition to the inhabitants.[4]

These plants have rich nutrition and medicinal values. Regular consumption of vegetables is also recommended for better health and management of chronic diseases. The nutritive value, antioxidant properties of the fruits of wild edible plants such as Bauhinia purpurea, Clerodendrum colebrookianum, Dillenia pentagyna, Diplazium esculentum, Houttuynia cordata, Oenanthe linearis, Potentilla lineata, Perilla ocimoides, Sonchus arvensis, and Zanthoxylum acanthopodium consumed by the tribal people of North-eastern region in India has already been studied in our laboratory. The quantification of phenolic acids and flavonoids by HPLC has also been carried out with these plants.

Therefore, these wild edible plants have nutritional potential and are worthy of exploitation as a dietary resource due to the presence of sufficient amount of protein, carbohydrate, fat, and minerals. The antioxidant properties and the presence of phenolic acids and flavonoids in these wild edible plants in varying amounts have enriched the nutraceutical properties of these plants.[5],[6],[7],[8],[9],[10],[11]

This paper accounts a simple, gradient, and stability-indicating HPLC method for the rapid determination of water-soluble vitamins such as thiamine (Vitamin B1), niacin (Vitamin B3), pyridoxine (Vitamin B6), ascorbic acid (Vitamin C), pantothenic acid (Vitamin B5), riboflavin (Vitamin B2), and folic acid (Vitamin B9) in ten wild edible plants named as B. purpurea, C. colebrookianum, D. pentagyna, D. esculentum, H. cordata, O. linearis, P. lineata, P. ocimoides, S. arvensis, and Z. acanthopodium from North-eastern region in India, and all the vitamins were simultaneously analyzed in a single chromatographic run.


   Materials and Methods Top


Plant material

The wild edible plants named B. purpurea, C. colebrookianum, D. pentagyna, D. esculentum, H. cordata, O. linearis, P. lineata, P. ocimoides, S. arvensis, and Z. acanthopodium were collected from North-eastern region in India. It was duly authenticated, and a voucher specimen was kept at the Department of Plant Chemistry of Botanical Survey of India under the Registry No. BSITS 15, BSITS 7, BSITS 16, BSITS 17, BSITS 3, BSITS 5, BSITS 10, BSITS 6, BSITS 4, and BSITS 1 for future reference. The plant parts were taken in our laboratory at refrigerated temperature using cold packs. The refrigerated plant samples were stored at −15°C and then processed within 4 days of collection.

Chemicals

The standards chemicals such as ascorbic acid (Vitamin C), thiamine (Vitamin B1), riboflavin (Vitamin B2), niacin (Vitamin B3), pantothenic acid (Vitamin B5), pyridoxine (Vitamin B6), and folic acid (Vitamin B9) were purchased from Sigma Chemical Co. (St. Louis, MO, USA), and the HPLC-grade solvents such as acetonitrile, methanol, water sodium dihydrogen phosphate, and trifluoroacetic acid were purchased from Merck (Germany).

High-performance liquid chromatography equipment

HPLC analyses were performed with Dionex Ultimate 3000 liquid chromatograph (Germany) with four solvent delivery system quaternary pump (LPG 3400 SD) including a diode array detector (DAD 3000) with 5 cm flow cell, a manual sample injection valve equipped with a 20 μl loop, and Chromeleon 6.8 system manager as data processor. The separation was achieved by a reversed-phase Acclaim™ 120 C18 column (5 μm particle size, i. d. 4.6 mm × 250 mm).

Preparation of standard solutions

The stock standard solutions of Vitamin C, B1, B3, B5, and B6 and were prepared by dissolving 25 mg of the each standard in one ml 0.1 M hydrochloric acid in 25 ml standard volumetric flask and topped up to mark with double distilled water. For preparation of standard stock solutions of Vitamin B9 and B2, 25 mg of the each standard were dissolved in one ml 0.1 M sodium hydroxide in 25 ml standard volumetric flask and made up to mark with double distilled water. The standard solution was stored in amber glass bottles in the refrigerator at 4°C. The working standards were prepared from the stock standard solutions by mixing 100 μl mixed vitamins standard (Vitamin B9, B5, and B2), 800 μl phosphate buffer (1 M, pH 5.5), and 100 μl mixed vitamins standard (Vitamin C, B1, B6, and B3) which represent 100 μg/ml mixed working standards. The working standard solutions of concentrations 20, 40, 60, and 80 μg/ml were prepared accordingly.

Preparation of sample solution

Plant materials were cleaned, and the inedible portions were removed. The edible parts were rinsed thoroughly with tap water and then with distilled water. The washed plant materials were dried with clean cloth, were cut into very small pieces, frozen in liquid nitrogen, freeze-dried, and kept at −20°C until analysis.

One g of each freeze-dried plant materials were soaked in 10 ml water. Then, 1 ml 0.1 M NaOH and 10 ml phosphate buffer (1M, pH 5.5) were added to it and kept in dark for 24 h. The solution was first filtered through a Whatman No. 1 filter paper and the resulting filtrate was taken in a 25 ml volumetric flask and solution was topped up to the mark with HPLC grade water. The sample solution was filtered through 0.45 μm membrane filter before injection into LC system. The stock solutions of sample were kept in a refrigerator for further use.[11]

Chromatographic analysis of water-soluble vitamins

The chromatographic analysis was carried out following the method as described by Marco Ciulua[12] with minor modification. The mobile phase contains acetonitrile (Solvent A) and aqueous trifluoroacetic acid (TFA, 0.01% v/v) (Solvent B), the column was thermostatically controlled at 22°C, and the injection volume was kept at 20 μl. A gradient elution was performed by varying the proportion of Solvent A to Solvent B. The gradient elusion was 1% solvent A and 99% solvent B with flow rate 0.5 ml/min in 5 min, from 1% to 25% solvent A with flow rate 0.5 ml/min for 16 min, from 25% to 45 % solvent A, with flow rate 0.5 ml/min for 8 min. from 45 to 1% solvent A with flow rate 0.5 ml/min in 5 min. The mobile phase composition back to initial condition (solvent A: solvent B : 1:99) in 34 min and allowed to run for another 1 min, before the injection of another sample. Total analysis time per sample was 35 min.

The various concentrations of (20, 40, 60, 80, and 100 μg/ml) vitamin working standards were injected into the HPLC column separately, and the retention times were noted and used to identify the vitamins in the sample.

HPLC chromatograms of all vitamins were detected using a photodiode array UV detector at four different wavelengths (210, 245, 275, and 290 nm) according to absorption maxima of analyzed compounds. Each compound in the plant extracts was identified by its retention time and by spiking with standards under the same conditions.

The quantification of the sample was done by the measurement of the integrated peak area, and the content was calculated using the calibration curve by plotting peak area against concentration of the respective standard sample. The data were reported as means ± standard error means of three independent analyses, and the method was validated according to the United States Pharmacopeia (USP) and International Conference on Harmonisation guidelines.[13],[14] Various parameters were studied to validate the reproducibility of the method, namely the effectiveness, the linearity, the limit of detection (LOD), the limit of quantitation (LOQ), the precision, and the accuracy.

Statistical analysis

Statistical analyses were performed according to the Statistical Package for Social Science (SPSS, version 22). The significant and nonsignificant variations within water-soluble vitamin contents and the ten wild edible plants were analyzed using one-way analysis of variance. The data were expressed as the mean ± standard error mean. P <0.05 was considered statistically significant.


   Results Top


Chromatographic method

A typical HPLC chromatogram of the all standard vitamin mixture recorded at 210 nm is presented in [Figure 1]. As shown in the chromatogram, all investigated compounds had responses at 245 nm, where they were successfully separated. The constituents under investigation were also identified by the recorded absorption spectra, which were comparable both for plant extracts and standard substances. The regression coefficient together with LOD and LOQ values is shown in [Table 1]. The high value of R2 > 0.9906 in the range of analyzed concentrations at 210, 245, and 275 nm is indicative of responsive linearity.
Figure 1: High-performance liquid chromatography chromatogram of mixture of standard vitamin. C: Ascorbic acid; B1: Thiamine; B3: Niacin; B6: Pyridoxine; B5: Pantothenic acid; B9: Folic acid; B2: Riboflavin

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Table 1: Retention time and parameters of calibration curve, precision and repeatability, limit of detection, limit of quantification, and percentage recovery study of standard water-soluble vitamins for high-performance liquid chromatography method validation

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Identification and quantification of water-soluble vitamins in the wild edible plants

The HPLC method was successfully performed for the estimation of water-soluble vitamin, for example, ascorbic acid (Vitamin C), thiamine (Vitamin B1), riboflavin (Vitamin B2), niacin (Vitamin B3), pantothenic acid (Vitamin B5), pyridoxine (Vitamin B6), and folic acid (Vitamin B9). The quantity of all vitamins of all plant materials has been expressed as mg/100 g dry plant material and data are presented in [Table 2].
Table 2: Quantification of Vitamin C and B1, B2, B3, B5, B6, and B9 in ten wild edible plants

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The HPLC analysis of the plants B. purpurea showed the presence of Vitamin C (15.35 ± 0.11 mg/100 g), Vitamin B1 (0.47 ± 0.01 mg/100 g), Vitamin B2 (0.047 ± 0.0001 mg/100 g), Vitamin B5 (0.32 ± 0.014 mg/100 g), Vitamin B6 (0.09 ± 0.003 mg/100 g), and Vitamin B9 (0.48 ± 0.0001 mg/100 g).

The HPLC study of the plant C. colebrookianum revealed the presence of Vitamin B2 (0.19 ± 0.002 mg/100 g), Vitamin B3 (0.02 ± 0.001 mg/100 g), Vitamin B5 (0.102 ± 0.002 mg/100 g), Vitamin B6 (0.18 ± 0.002 mg/100 g), and Vitamin B9 (0.31 ± 0.002 mg/100 g).

The presence of Vitamin B1 (0.02 ± 0.0005 mg/100 g), Vitamin B2 (0.08 ± 0.001 mg/100 g), Vitamin B3 (0.37 ± 0.001 mg/100 g), Vitamin B5 (0.08 ± 0.005 mg/100 g), Vitamin B9 (0.13 ± 0.002 mg/100 g), and remarkable amount of Vitamin B6 (0.82 ± 0.004 mg/100 g) were detected in D. pentagyna.

The leaves of D. esculentum were found to contain Vitamin C (5.41 ± 0.03 mg/100 g) along with Vitamin B1 (0.006 ± 0.0005 mg/100 g), Vitamin B2 (0.09 ± 0.001 mg/100 g), Vitamin B5 (0.07 ± 0.001 mg/100 g), Vitamin B6 (0.12 ± 0.001 mg/100 g), and significant amount of Vitamin B9 (1.31 ± 0.001 mg/100 g).

The HPLC analysis showed the presence of Vitamin B1 (0.0004 ± 0.00003 mg/100 g), Vitamin B2 (0.03 ± 0.001 mg/100 g), Vitamin B3 (0.006 ± 0.0002 mg/100 g), Vitamin B5 (0.40 ± 0.003 mg/100 g), Vitamin B6 (0.04 ± 0.0002 mg/100 g), and good amount of Vitamin B9 (0.42 ± 0.002 mg/100 g) in the roots of H. cordata.

Our investigation disclosed the occurrence of Vitamin B1 (0.0061 ± 0.002 mg/100 g), Vitamin B2 (0.057 ± 0.003 mg/100 g), Vitamin B3 (0.027 ± 0.001 mg/100 g), Vitamin B5 (0.037 ± 0.001 mg/100 g), Vitamin B6 (0.10 ± 0.002 mg/100 g), and Vitamin B9 (0.56 ± 0.002 mg/100 g) in the leaves of O. linearis.

The HPLC study of the seeds of P. ocimoides showed the presence of Vitamin B2 (0.20 ± 0.003 mg/100 g), Vitamin B3 (0.012 ± 0.001 mg/100 g), Vitamin B5 (0.16 ± 0.007 mg/100 g), and Vitamin B6 (0.08 ± 0.001 mg/100 g) along with very sufficient amount of Vitamin B9 (0.33 ± 0.003 mg/100 g).

The tuberous roots of P. lineata were found to contain Vitamin B2, Vitamin B6, and Vitamin B9 amounting 0.29 ± 0.002, 0.24 ± 0.005, and 0.30 ± 0.004 mg/100 g, respectively, whereas Vitamin C, B1, B3, and B5 were not detected.

The HPLC study of the leaves of S. arvensis showed the presence of Vitamin B2 (0.18 ± 0.002 mg/100 g), Vitamin B5 (0.27 ± 0.004 mg/100 g), Vitamin B6 (0.36 ± 0.004 mg/100 g), and Vitamin B9 (0.10 ± 0.004 mg/100 g) whereas the leaves of Z. acanthopodium found to contain vitamins (C, B1, B2, B5, B6, and B9) amounting 0.76 ± 0.03,0.001 ± 0.00002, 0.38 ± 0.001, 0.05 ± 0.001, 0.04 ± 0.001, and 0.40 ± 0.001 mg/100 g, respectively.


   Discussion Top


Chromatographic method

The quantitative analysis of water-soluble vitamins was carried out using a photodiode array UV detector at four different wavelengths (210, 245, 275, and 290 nm). The detection of Vitamin C, Vitamin B1, and Vitamin B3 were carried out at wavelength 245 nm, Vitamin B2, Vitamin B6, and Vitamin B9 were carried out at 275 nm. The detection wavelength was set at 210 nm for Vitamin B5 as it showed absorption at 209 nm. The chromatographic separation was performed at a flow rate of 0.5 ml/min. The method proposed was rapid, and all analytes were completely eluted within 30 min, and the whole chromatographic run was completed in 35 min. The solvent system (acetonitrile and aqueous TFA, 0.01% v/v) was used for the analysis and produced a sharp peak of the studied vitamins.

The repeatability of the retention time for all the standard samples and the relative standard deviation for the peak areas of two standards, namely 40 and 60 μg/ml was found to be below 1%. In this chromatographic method the percentage of recovery of standard vitamins is significantly high (98.15–99.20%). It follows that the method under consideration is characterized by precision, accuracy, and meticulousness and can be used for the qualitative as also quantitative estimation of water-soluble vitamins in the ten wild edible plants under investigation.

The aim of this study was to develop simple, gradient, and stability-indicating HPLC method for the determination of Vitamin C, B1, B2, B3, B5, B6, and B9 in ten wild edible plants. Vitamin C is extremely unstable in basic and neutral solutions, but relatively stable in acidic solutions; therefore, phosphate buffer (pH 5.5) was used as a diluting solution for Vitamin C, B1, B3, B5, and B6. Both the Vitamins (B2 and B9) were found slightly soluble in water and acidic aqueous solutions, but soluble in basic aqueous solutions. Hence, the stock solutions of Vitamin B2 and B9 were dissolved in 0.1M NaOH solution, and all working standard vitamins were diluted with phosphate buffer (pH 5.5) solution.

Identification and quantification of water-soluble vitamins in the wild edible plants

Vitamin C is the most important vitamin in fruits and vegetables. Plants, rich in Vitamin C, are well known for its antioxidant properties, and they help the body in inhibiting from viral infection, bacterial infections, and toxicity. It is required for the prevention of scurvy and maintenance of healthy skin, gums, and blood vessels, and the deficiency of this vitamin causes bruising, bleeding, dry skin, and depression.[15],[16]

The experimental result showed that the amount of Vitamin C in ten wild edible plants ranged from 0.76 ± 0.03–15.35 ± 0.11 mg/100 g. The quantity of Vitamin C was found highest in the leaves of B. purpurea (95.54 ± 3.33 mg/100 g) followed by in the leaves of D. esculentum (5.41 ± 0.03 mg/100 g) [Figure 2]. The Vitamin C content in these wild edible plants is very much comparable with some common fruits and vegetables such as Solanum tuberosum (17.04 ± 1.18 mg/100 g), Allium sativum (13.06 ± 1.10 mg/100 g), and Daucus carota sativus (2.55 ± 0.72 mg/100 g).[15] Vitamin C was not detected in other plants under investigation.
Figure 2: Vitamin C content in ten wild edible plants

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Hence, the wild edible plants under investigation might be considered good sources of Vitamin C and therefore could be able to satisfy the recommended daily allowances of 75 mg/day and 90 mg/day for adult women and men, respectively, and 45 mg/day for children of 9–12 years old. Due to having antioxidant properties, Vitamin C-rich plant might be beneficial to reduce the risk of atherosclerosis and some forms of cancer.[17]

Thiamine (Vitamin B1) is an essential nutrient required by the body for maintaining cellular function and consequently a wide array of organ functions. It is indispensable for energy production, carbohydrate metabolism, and nerve cell function. The deficiency of this vitamin is responsible for high blood pressure, cardiac diseases and deterioration of the nervous systems.[18],[19]

The thiamine content in these wild edible fruits ranged from 0.0004 ± 0.00001 to 0.47 ± 0.01 mg/100 g. The highest amount of Vitamin B1 was obtained from the leaves of B. purpurea followed by D. pentagyna and D. esculentum. The sufficient amount of this vitamin was also noted in H. cordata, O. linearis, and Z. acanthopodium and was not detected in other four plants under study [Figure 3].
Figure 3: Vitamin B1, B2, and B3 content in ten wild edible plants

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Thiamine has been shown to occur in some common vegetables and fruits such as apple (0.016 mg/100 g), beans (0.132 mg/100 g), cauliflower (0.073 mg/100 g), and spinach (0.076 mg/100 g), and these amounts are very much similar to the thiamine content detected in the wild edible plants under investigation.

Riboflavin (Vitamin B2) is a vital vitamin required for proper energy metabolism and a wide variety of cellular processes. It is the counterpart to thiamine used in the strengthening of food products.[20] A significant variation of riboflavin content was noticed among the tested wild edible plants. The highest amount of Vitamin B2 was detected in the leaves of Z. acanthopodium (0.38 ± 0.001 mg/100 g) and the least amount was detected in H. cordata (0.03 ± 0.001 mg/100 g). The leaves of C. colebrookianum, S. arvensis, seeds of P. ocimoides, and tuberous roots of P. lineata were also found to contain a very good quantity of Vitamin B2[Figure 3] which are comparable with some common fruits and vegetables such as almonds (1.10 mg/100 g), spinach (0.24 mg/100 g), beet greens (0.41 mg/100 g), green beans (0.12 ± 2 mg/100 g), and potato (0.023 ± 1 mg/100 g).[21]

The niacin (Vitamin B3) content in the wild edible plants under analysis ranged between 0.006 ± 0.0002 mg/100 g and 0.37 ± 0.001 mg/100 g. The highest amount of Vitamin B3 was detected in D. pentagyna followed by in the leaves of O. linearis (0.027 ± 0.001 mg/100 g) [Figure 3]. Therefore, these plants are the important sources of Vitamin B3. The other plants, namely C. colebrookianum, H. cordata, and P. ocimoides also found to have necessary amount of Vitamin B3 which were comparable with cabbage, cauliflowers, cucumber, spinach, and tomatoes ranged between 0.19 and 0.97 mg/100 g.[22]

Vitamin B3 is an important vitamin required for processing fat in the body, lowering cholesterol levels, and regulating blood sugar levels. It is important in DNA repair, Ca metabolism, intracellular respiration, and biosynthesis of fatty acid and steroids.[23] Hence, the regular consumption of these plants would supply adequate Vitamin B3 necessary to maintain healthy body functions.

Vitamin B5, or Pantothenic acid, is an essential vitamin required by the body for cellular processes and optimal maintenance of fat. The deficiency of Vitamin B5 leads to irritability, fatigue, apathy, numbness, paresthesia, and muscle cramps in human being.[24]

Pantothenic acid was detected highest in the roots of H. cordata (0.40 ± 0.003 mg/100 g). The leaves of B. purpurea, C. colebrookianum, and S. arvensis were also found to contain a very good amount of Vitamin B5 and was not detected in P. lineata [Figure 4].
Figure 4: Vitamin B5, B6, and B9 content in ten wild edible plants

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Pyridoxine (B6) is another water-soluble vitamin necessary for the proper maintenance of red blood cell metabolism, the nervous system, the immune system, and many other bodily functions. It also plays a role in homocysteine synthetic and degradative reactions.[25] This vitamin is found in most food products and also, due to its stability, is often used for fortifying food products.[26] It was quantified in all the wild edible plants under our investigation. The highest Vitamin B6 was observed in D. pentagyna (0.82 ± 0.004 mg/100 g) whereas the minimum was detected in both H. cordata and Z. acanthopodium (0.04 mg/100 g). The Vitamin B6 content ranged between 0.08 ± 0.001 and 0.36 ± 0.004 mg/100 g in other seven wild edible plants under investigation [Figure 4].

The amount of Vitamin B6 obtained in these wild edible plants was comparable with some common vegetable and fruits such as banana (0.37 mg/100 g), avocados (0.29 mg/100 g), spinach (0.24 mg/100 g), broccoli (0.134 mg/100 g), cauliflower (0.115 mg/100 g), and cucumber (0.2 mg/100 g). Hence, the regular intake of these plants would supply sufficient Vitamin B6 necessary to maintain healthy body functions.

Vitamin B9(folic acid) is a water-soluble B vitamin with many rich natural sources. It is required for numerous body functions including DNA synthesis and repair, cell division, and cell growth. The shortage of folate can lead to anemia in adults and slower development in children.[27],[28],[29],[30] It plays an important role as an antioxidant in vivo, both by preventing the adverse effect of reactive oxygen species), as well as by inhibiting lipid peroxidation.[30]

The extent of Vitamin B9 in ten wild edible plants ranged from 0.13 ± 0.001 to 1.31 ± 0.002 mg/100 g. The content of Vitamin B9 was found highest in D. esculentum and the leaves of O. linearis contained second highest amount of Vitamin B9 (0.56 ± 0.002 mg/100 g). Among ten wild edible plants, Vitamin B9 was lowest in D. pentagyna. A good amount of Vitamin B9 was also detected in the leaves of B. purpurea (0.48 ± 0.0001 mg/100 g), S. arvensis (0.60 ± 0.004 mg/100 g), and Z. acanthopodium (0.40 ± 0.001 mg/100 g) [Figure 4].


   Conclusion Top


The reversed-phase HPLC method with diode array detection was developed for the quantitative estimation of water-soluble B vitamins (B1, B2, B3, B5, B6, and B9) and Vitamin C in ten wild edible plants such as B. purpurea, C. colebrookianum, D. pentagyna, D. esculentum, H. cordata, O. linearis, P. lineata, P. ocimoides, S. arvensis, and Z. acanthopodium collected from North-eastern region in India. The established HPLC assay showed a well separation of the compounds, and also, the developed method was linear, sensitive, accurate, meticulous, and reproducible. Therefore, the method can be used for the simultaneous determination of water-soluble B vitamins and Vitamin C in different formulations with “shorter run time” and “high efficiency.” RP-HPLC results showed that the plants contained several water-soluble B and C vitamins in varying amounts. The result of analysis of vitamin content in the wild edible plants under investigation will serve as a useful means to calculate dietary intake of C and B vitamins in the general population. These data will also be helpful in the preparation of a complete food composition table for nutritional survey and also for other research purposes.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4]
 
 
    Tables

  [Table 1], [Table 2]



 

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