Introduction

Green leafy vegetables (GLV) are multi-cultural components used ubiquitously in Indian cuisine. They are rich sources of calcium, iron, β-carotene, vitamin C, dietary fiber and many trace minerals. A large number of leaves from different sources such as perennial trees, aquatic plants and annuals are consumed and especially in rural areas. These vegetables are an economic source to ensure the micronutrient intake. GLV are seasonal and also highly perishable due to their high water content. There are heavy losses due to non-availability of sufficient storage, transport and proper processing facilities at the production point (Pande et al. 2000). There is a need to preserve the nature’s storehouse of nutrients through convenient processing techniques. Dehydration seems to be the simplest technology for preserving GLV, especially when they are abundantly available.

Vegetable dehydration is generally done either for preserving the perishable raw commodity against deterioration or to reduce the cost of packaging, handling, storing and transporting. The most serious constraint for shelf-life enhancement is the activity of micro-organisms. Water in food is reduced to a very low level during dehydration, thus achieving better microbiological preservation and retarding many undesirable reactions during storage (Ibarz and Barbosa-Canovas 2000), owing to the reduction in water activity. In the present study dehydration technology is utilized with a different perspective. GLV are rich sources of micronutrients as already discussed earlier and therefore dehydrating them can provide us a concentrated source of micronutrients. Utilizing these micronutrient-rich GLV in a dehydrated form can be a food-based approach to combat the micronutrient deficiencies, which is prevalent in our populations, especially during seasons of their non-availability.

In recent years, exhaustive efforts have been made for an improvement in the nutrient retention of dried products by altering processing methods and/or pretreatment. Blanching is a prerequisite for preservation of green leafy vegetables. It is necessary to prevent the formation of off-flavors, odors and colors. However, it may cause partial destruction of some nutrients like ascorbic acid. Peroxidase activity is widely used as an index of blanching because peroxidase is the most heat-stable enzyme found in vegetables (Rahman and Perera 1999). Optimum conditions of blanching, time and temperature, are necessary to achieve the desired quality of dried products (Kadam et al. 2008). The protective effect of sodium metabisulphite has been demonstrated by Mulay et al. (1994) and Badifu et al. (1995), which is supported by Singh et al. (1997). The latter reported that the use of potassium metabisulphite (KMS) for pretreatment of the green leafy vegetables can reduce the extent of loss of ascorbic acid. Blanching also results in some degree of chlorophyll degradation with the subsequent formation of pheophytin, which can be prevented by the addition of an alkalizing agent like magnesium carbonate during blanching (Maharaj and Sankat 1996). The extent of chlorophyll conversion is related to the degree of blanching. A combination of chemicals (0.5% potassium metabisulphite + 0.1% magnesium oxide + 0.1% sodium bicarbonate) used for blanching is known to have better retention of ascorbic acid when compared to individual chemicals used for blanching (Patil et al. 1978, Premavalli et al. 2001). Negi and Roy (2000) have reported that water blanching followed by dipping in potassium metabisulphite and drying amaranth and fenugreek in a low temperature drier had the least drastic effect on β-carotene, ascorbic acid and chlorophyll content of the processed product.

The present investigation was therefore undertaken to evaluate the influence of dehydration on the nutritional composition of GLV and to produce a concentrated source of micronutrients in the form of dried greens.

Materials and methods

Selection and preprocessing of GLV

Five GLV, namely Amaranth leaves (Amaranthus gangeticus), Brahmi (Centella asiatica), Bathua leaves (Chenopodium album) Kilkeerae (Amaranthus tricolor) and Fenugreek leaves (Trigonella foenum graecum) were selected to study the changes in composition on dehydration. The fresh GLV were procured from local markets. The leaves were separated from the roots and washed under running water to remove the adhering mud particles followed by double glass-distilled water and drained completely. One set of greens was used as such for analysis. Another set of GLV was steam blanched for 5 min after chemical treatment with a solution of 0.1% magnesium oxide + 0.1% sodium bicarbonate and 0.5% potassium metabisulphite. They were drained after blanching and spread on steel trays for drying in an oven at 60°C for 10–12 h. After drying the GLV were powdered in a mixer, stored in an airtight container and stored in a refrigerator.

Compositional analysis

The fresh GLV and dehydrated GLV were analyzed for the following components to study the effect of dehydration. Moisture, ether extractives and ash (minerals) were estimated by standard methods (AOAC 2000). Total iron was analyzed by colorimetric method using α – α bipyridyl (AOAC 1965) and calcium by the method of AOAC (2000). Insoluble and soluble dietary fiber was analyzed by separation of non-starch polysaccharides by the enzymatic gravimetric method (Asp et al. 1983). Ascorbic acid was estimated by the visual titration method of reduction of 2, 6—dicholorophenol-indophenol dye. Total carotene was extracted in acetone; β-carotene was separated by column chromatography and estimated colorimetrically (Ranganna 1986). Thiamine was analyzed by oxidation to thiochrome, which fluoresces in UV light (Raghuramulu et al. 2003). Total oxalate was analyzed by extraction with hydrochloric acid and soluble oxalate with water followed by precipitation with calcium oxalate from deproteinized extract and subsequent titration with potassium permanganate (Gupta et al. 2005). Tannins were extracted in methanol and measured colorimetrically by using vanillin-hydrochloride method (Gupta et al. 2005).

Analysis of in vitro dialyzability of Fe and Ca

In vitro dialyzability of iron and calcium from GLV was determined by simulated gastrointestinal digestion using pepsin for the gastric stage followed by pancreatin and bile salts for the intestinal stage (Luten et al. 1996). The proportion of mineral that diffused through a semipermeable membrane was used to measure mineral dialyzability. Exactly 15 g of homogenized green leafy vegetables (5 g in case of dried samples) were placed in a 250 ml Erlenmeyer flask and 80 ml of water was added. The pH was adjusted to 2.0 with 6 M HCl. After 15 min, the pH was checked and readjusted to 2.0, if necessary. Exactly 3 ml of freshly prepared pepsin solution (16% in 0.1 M HCl) was added and the volume was made up to 100 ml with water. The samples were incubated at 37°C for 2 h. The digests were then frozen for 90 min. Titratable acidity was determined in an aliquot of the digest containing 5 ml of pancreatin-bile extract mixture (0.4% pancreatin + 2.5% bile extract) with 0.2 M NaOH till the pH of 7.5 was attained.

Exactly 20 ml of frozen digests (after thawing) were subjected to simulated intestinal digestion by placing the dialysis tubing (molecular cut off: 8 Kda) in Erlenmeyer flasks. The dialysis tubing contained 25 ml of NaHCO3 (equivalent to moles of NaOH determined by titratable acidity) solution. The flasks were incubated in a shaker water bath at 37°C for 30 min (until the pH reached 5.0), 5 ml of pancreatin - bile extract mixture was added and shaken for another 2 h (until pH reached 7.0). Then the dialysates were carefully transferred to graduated tubes and the volume was measured. The dialysates were analyzed for iron by colorimetric method using α - α bipyridyl (AOAC 1965) and calcium by the method of Raghuramulu et al. (2003). Double glass-distilled water was used for preparation of reagents used in the entire analysis. All chemicals used for the study were of analytical grade.

Statistical analysis

The data presented represents the mean of quadruplicate analysis, standard deviation was computed to assess the variation between the replicate values. ANOVA and the students ‘t’ tests were suitably applied to study the compositional differences between the fresh and blanched dehydrated samples. Probability level was set to P < 0.05. The statistical package used for the above analysis was Minitab 1.32.

Results and discussion

In the present study we have analyzed the effect of dehydration on the nutritive value of GLV. The results of the study are presented in Tables 1, 2, 3, 4 and Fig. 1.

Table 1 Proximate and mineral composition of fresh and dehydrated green leafy vegetables
Full size table
Table 2 Vitamin content of fresh and dehydrated green leafy vegetables* (mg/100 g)
Full size table
Table 3 Antinutrient content of fresh and dehydrated green leafy vegetables (mg/100 g)
Full size table
Table 4 Dialyzable iron and calcium from fresh and dehydrated green leafy vegetables (mg/100 g)
Full size table
Fig. 1
figure 1

Percent dialyzable iron and calcium from fresh and dehydrated green leafy vegetables

Full size image

The moisture content of the fresh GLV ranged between 85.7–92.2% (Table 1). On dehydration, the moisture content was found to be in the range of 3.5–7.9%. Drying of GLV seems to have no effect on the ether extract which is evident from the results (Table 1). No significant differences were observed between the ether extact of fresh and dehydrated GLV when t-test was applied to check for differences (P = 0.119ns). When the ash content of the fresh and dehydrated GLV was considered, the maximum amounts were found in Amarnathus tricolor in both the fresh and dry forms. In the other GLV, it was found to be in the range of 1.36–1.88 g/100 g in fresh and 1.28–2.02 g/100 g of dry vegetables. As per the student’s t-test, no significant differences were observed in the ash content on dehydration (P = 0.416ns).

The effect of dehydration on the retention of iron and calcium was studied. The results are presented in Table 1. Among the fresh GLV, Amaranthus tricolor had the maximum amount of iron, whereas Chenopodium album had the least. Amaranthus gangeticus, Centella asiatica and Trigonella feonum graecum had iron contents of 16.38, 12.46 and 4.05 mg/100 g respectively. Dehydration does not seem to have much effect on the iron content of the GLV. Although there were variations in the iron content of dehydrated GLV when compared to the fresh, no significant differences were seen on application of the t-test (P = 0.181ns). The calcium content of the fresh and dehydrated GLV was found to be similar. The slight variations that were observed were found to be statistically insignificant (P = 0.483ns). Not many studies have reported the effect of dehydration on mineral content of GLV although Kawashima and Valente Soares (2003) have reported that brief cooking did not cause appreciable losses of any of the minerals in leafy vegetables consumed in South Brazil.

When we consider the insoluble dietary fiber content of the fresh GLV, it was in the range of 1.86 g/100 g in Chenopodium album to 5.08 g/100 g in Centella asiatica (Table 1). In the other GLV, it was in the range of 2.99–3.87 g/100 g. In all the GLV, a decrease in the insoluble fiber content was seen on dehydration. These differences on the insoluble dietary fiber content in the fresh and dehydrated GLV were found to be statistically insignificant according to the student’s t-test (P = 0.284ns).

A different trend was seen in the soluble dietary fiber content of the leafy vegetables. In the fresh samples the soluble fiber was found to be in the range of 0.32–0.80 g/100 g (fresh weight basis). On dehydration, a marginal increase was seen in Chenopodium album and Centella asiatica. In the rest of the samples, a marginal decrease in the soluble dietary fiber content was seen on dehydration. The statistical analysis revealed the differences to be insignificant (P = 0.462ns). There seems to be no trend of increase or decrease in the dietary fiber content of the GLV. These differences could only be attributed to the variety of samples that were used for the study. Punna and Rao (2004) have reported no significant effect of processing/cooking on insoluble and soluble dietary fiber contents of green leafy vegetables.

Ascorbic acid, thiamine, total and β-carotene were the vitamins that were analyzed in both fresh and dehydrated GLV. Ascorbic acid, the most water-soluble and heat labile vitamin was found to be lost in large quantities on dehydration. As can be seen in Table 2, the fresh GLV have good amounts of ascorbic acid ranging between 13.8–99.7 mg/100 g fresh vegetable. On dehydration, the ascorbic acid decreased to a range of 0.5–8.2 mg/100 g (fresh weight basis). Reduction in the ascorbic acid content on dehydration was found to be statistically significant (P = 0.005*). Ascorbic acid is a vitamin that is lost very easily. When we consider the amount of ascorbic acid that was retained on dehydration, a considerable variation was seen. Almost all ascorbic acid was destroyed in Chenopodium album whereas Centella asiatica retained 14% of its initial ascorbic acid content. Among the other GLV, the retention of ascorbic acid was in the range of 4–8%. Conversely Kaur et al. (2008) have reported the retention of ascorbic acid in mustard, mint and spinach to be as high as 13–38%.

Fresh Trigonella foenum graecum was found to have the maximum amounts of thiamine, followed by Amaranthus gangeticus and Centella asiatica. Fresh Chenopodium album and Amaranthus tricolor had similar amounts of thiamine. Dehydrated Trigonella foenum graecum had 0.30 mg/100 g (fresh weight basis) followed by Centella asiatica and Amaranthus gangeticus (0.06, and 0.04 mg/100 g respectively). Dehydration did result in losses of thiamine. Statistical analysis showed no significant differences between the thiamine content of fresh and dehydrated GLV inspite of the losses on dehydration (P = 0.184ns). In comparison with ascorbic acid, thiamine was better retained on dehydration. Almost 71% retention was seen in Trigonella foenum graecum, followed by Amaranthus tricolor and Centella asiatica, which retained 48 and 43% of thiamine on dehydration. Amaranthus gangeticus and Chenopodium album retained 22 and 31% of their initial thiamine content after drying.

The total carotene content of the GLV was also affected by dehydration. Among the GLV selected for the study, Centella asiatica had the maximum amounts of total carotene in the fresh sample, which decreased by 27% on dehydration. The total carotene content decreased by 36.6% in Amaranthus tricolor, 52.9% in Trigonella foenum graecum, 31.9% in Amaranthus gangeticus and 45.4% in Chenopodium album on dehydration. Statistically significant differences were observed in total carotene on dehydration (P = 0.018*). β-carotene, which is a precursor of vitamin A was also analyzed. Among the GLV, the least amount of β-carotene was found in Chenopodium album that decreased by 79.6% on drying. The rest of the GLV had β-carotene in the range of 4.35–5.46 mg/100 g that reduced to 2.69–3.63 mg/100 g on dehydration. Dehydration losses were found to be statistically significant (P = 0.020*). These results are in agreement with Kaur et al. (2008) who have studied the retention of β-carotene in dehydrated mustard, mint and spinach greens. Uadal and Sagar (2008) have reported that dehydrated amaranth, fenugreek and spinach retained higher β-carotene and ascorbic acid when stored in high density polyethylene at low temperature. From the above results, it can be said that vitamins are more prone to destruction on dehydration, while there seems to be little effect on the other proximate constituents.

Tannins, total and soluble oxalates were analyzed in fresh and dehydrated GLV. In the fresh samples, the tannin content (expressed as catechin equivalents) was found to be the least in Chenopodium album and highest in Amaranthus tricolor. Among the other GLV, it ranged between 122.5–170.9 mg/100 g (Table 3). On dehydration, no trend was seen in the tannin content of the samples. This could be attributed to the sample itself. Differences in the tannin content of fresh and dehydrated GLV were found to be statistically insignificant (P = 0.189ns).

The total oxalate content of Amaranthus gangeticus and Trigonella foenum graecum was found to increase on dehydration, whilst in the other GLV the total oxalate content was found to decrease on dehydration. In the fresh and dehydrated samples, total oxalate ranged between 78.3–1243.3 and 18.2–1216.9 mg/100 g, respectively. No significant differences were seen in the total oxalate content as a result of dehydration (P = 0.479ns). A similar trend was seen in the soluble oxalate content of the dehydrated GLV. The soluble oxalate was seen to increase in Amaranthus gangeticus and Trigonella foenum graecum on dehydration, whilst in the other GLV dehydration seemed to have reduced the soluble oxalate content. The soluble oxalate was found to be in the range of 8.7–719.6 and 9.1–414.4 mg/100 g of fresh and dehydrated GLV, respectively. The differences between the soluble oxalate content of fresh and dehydrated GLV were found to be statistically insignificant (P = 0.268ns).

There are very few/limited studies on the effect of dehydration on the antinutrient content of the green leafy vegetables in the literature. However some literature studies have reported that cooking/conventional boiling method was effective in reducing the oxalic acid and tannin content of some GLV (Savage et al. 2000, Somsub et al. 2008). From the above observations, it can be concluded that dehydration seems to have little effect on the proximate, mineral and antinutrient contents of the GLV. However, statistically significant differences were seen only in the vitamin content of the GLV on dehydration.

Experiments were also carried out to analyze the effect of dehydration on dialyzable iron and calcium by in vitro technique. From Table 4, it can be seen that the dialyzable iron in the fresh GLV was found to be maximum in Amaranthus tricolor followed by Centella asiatica, which had 3.50 mg/100 g. In the rest of the fresh samples, the dialyzable iron was found to be in the range of 0.21–0.65 mg/100 g, respectively. When dehydrated GLV were considered, we found a sharp decline in the dialyzable iron of all the dried greens with the exception of Amaranthus gangeticus, which had higher dialyzable iron content in the dehydrated form than in the fresh form. The maximum amount of absolute dialyzable iron in the dried greens was seen in Amaranthus tricolor. In the remaining dehydrated greens the dialyzable iron was found to be in the range of 0.05 mg/100 g in Trigonella foenum graecum to 0.50 mg/100 g in Centella asiatica.

When we consider the percent dialyzable iron, in the fresh GLV it was in the range of 2.4–31.4%, while in the dehydrated greens it was found to be in the range of 2.7–11.1% (Fig. 1). Different workers have reported varying values for percent dialyzability from green leafy vegetables. Chawla et al. (1988) in one of the earliest studies determined the in vitro availability of iron and related constituents in six green leafy vegetables (amaranth, colocasia, drumstick, fenugreek, shepu and spinach) and found it to be between 2.8–4.6%. Lucarini et al. (2000) studied the dialyzable iron content from artichoke, asparagus, broccoli, cabbage, cauliflower, kale, carrot, tomato and potato. The in vitro dialyzable iron ranged between 10.7–23.1% with the exception of artichoke and asparagus. These findings have also been reported by Gillooly et al. (1983). The in vitro iron bioavailability in the uncommon GLV from the Uttaranchal region of India ranged between 4.62–6.20% (Raghuvanshi et al. 2001). These differences in the dialyzable iron content of the vegetables can be attributed to the variations in the organic acid content of vegetables and the presence of the antinutrients in these vegetables. To the best of our knowledge this could be the first report to show the effect of dehydration on the dialyzable iron content of GLV. The decrease in dialyzability on dehydration could be attributed to the fact that during the process of dehydration, iron could have been bound to other constituents of the vegetables thus reducing the solubility, which in turn influences the dialyzability of the mineral. Also, ascorbic acid that is a promoter of iron bioavailability is also destroyed during dehydration.

The absolute amount of dialyzable calcium was found to be highest in Centella asiatica and Trigoenlla foenum graecum in the fresh samples. In the other GLV, it ranged from 15.36–27.88 mg/100 g of fresh sample (Table 4). When the dehydrated GLV were considered, we found a decline in their dialyzable calcium content with respect to the fresh greens. In Chenopodium album the dialyzable calcium reduced by more than half from 27.88 mg/100 g of fresh greens to 6.94 mg/100 g of dried greens. Similarly in Centella asiatica and Trigonella foenum graecum the dialyzable calcium content reduced from 81.33 to 58.15 and 61.25 to 39.96 mg/100 g, respectively, on dehydration.

Chenopodium album and Centella asiatica had the highest calcium dialyzability of 88.3 and 42.1% in the fresh GLV. On dehydration, the calcium dialyzability decreased to 17.4 and 35.5% respectively. In Amaranthus gangeticus, Amaranthus tricolor and Trigonella foenum graecum, the bioavailable calcium reduced from 8.3 to 6.1%, 9.1 to 3.9% and 39.4 to 27.3% (Fig. 1). Lucarini et al. (1999) evaluated the in vitro calcium availability from four varieties of Brassica oleracea L. (broccoli, cauliflower, green cabbage and kale). Twenty five percent of the total calcium was found to be dialyzable. Since Brassica vegetables are essentially phytate- and oxalate-free vegetables, dietary fiber components and organic acids could have influenced calcium availability. Raghuvanshi et al. (2001) reported that inspite of high oxalate content of uncommon GLV from Uttaranchal regions of India, the in vitro calcium bioavailability ranged between 7.30–63.48%. Conversely, Kamchan et al. (2004) reported high levels of dialyzable calcium (20–39%) in kale, celery, collard, pak-chee-lao (Anethum graveolens, L.), Chinese cabbage and soybean sprouts. Medium levels of dialyzable calcium (11–18%) were found in Indian mulberry and sesbania leaves. Pak-paw (Polygonum odoratum, L), amaranth and wild betel exhibited low calcium availability. The differences in the calcium bioavailability as reported by Raghuvanshi et al. (2001) and Kamchan et al. (2004) could be attributed to the different methods used for evaluation of in vitro available calcium.

A large variety of interactive food components were found to interfere in one or other form with the bioavailability of iron and calcium. The availability of iron and calcium in fresh GLV was found to be in the range of 2.4–31.4% and 8.3–88.3%, respectively, and that in the dehydrated GLV in the range of 2.7–11.1% and 3.9–40.0%, respectively. In our study, the inhibitors influenced the availability significantly and dehydration seems to have a negative influence on bioavailability of iron and calcium in GLV. Recently, some studies have reported that the disruption of the natural matrix or the microstructure created during processing may influence the release, transformation, and subsequent absorption of some nutrients in the digestive tract. Parada and Aguilera (2007) have reviewed recent scientific data that demonstrate that in the case of certain nutrients, the state of the matrix of natural foods or the microstructure of processed foods may favor or hinder their nutritional response in vivo.

Conclusion

The compositional changes that occurred on dehydration varied by the component. Proximate principles were least affected, total iron and calcium content decreased slightly, but dialysability of the minerals decreased significantly. Among the vitamins, ascorbic acid, total and β- carotene were lost significantly, while thiamine was retained moderately. Changes in the antinutritional factors was non-significant. The process of dehydrating GLV concentrates the nutrients and the dehydrated GLV are a rich source of dietary fiber which can find application in development of high fiber and micronutrient rich foods. Easy to preserve, feasibility, convenience and offseason availability are some of the advantages of incorporating dehydrated GLV in products. Dehydrated GLV can be incorporated into traditional products at the household level or can be utilized in the formulation of processed foods at the industrial level. Value addition of food products with dehydrated GLV can be advocated as a feasible food-based approach to combat micronutrient malnutrition.