Evaluation of mineral composition and in-vitro nutrient digestibility of macrophytes to assess their potential as sustainable animal feed

Abstract

The current study evaluates the macro- and micro-mineral profiles, assesses heavy metal concentrations, and determines the in-vitro digestibility of various aquatic macrophytes collected from Dal Lake, Manasbal Lake, Hokersar Lake, and Anchar Lake. The macro-mineral analysis reveals significant variations among different species in each lake, highlighting the nutritional diversity of these aquatic plants. Lemna minor from Dal Lake, Azolla cristata from Manasbal Lake, Rumex rupestris from Hokersar Lake, and Lemna minor and Azolla cristata from Anchar Lake exhibited noteworthy macro-mineral concentrations. In terms of micro-minerals, Copper (Cu) concentrations were consistent among macrophytes from Dal Lake, while Fe levels were significantly higher (p < 0.05) in Nymphaea tetragona. The study found similar trends in micro-mineral concentrations in macrophytes from other lakes. The heavy metal analysis demonstrated varying concentrations among macrophytes in Dal Lake and Manasbal Lake, with some species showing potential as phytoremediators. The outcomes of in-vitro digestibility revealed significantly higher (p < 0.05) digestibilities of dry matter (DM), organic matter (OM), and neutral detergent fiber (NDF) in weeds Nelumbo nucifera, Trapa natans, and Lemna minor sourced from Dal Lake, whereas, Nymphaea tetragona from Manasbal Lake also revealed significantly higher values (p < 0.05). Significantly lower (p < 0.05) in-vitro digestibility values were revealed by Typha angustata, Nymphoides aquatica, and Ceratophyllum demersum from Nigeen Lake, likewise Nymphoides peltata sourced from Anchar Lake exhibited lower values. The study emphasizes the impact of environmental factors on mineral accumulation and the potential use of aquatic plants for nutrient removal and phytoremediation.

Introduction

Aquatic macrophytes are fastest growing taxonomically diverse plants whose life cycle takes place completely or periodically in the aquatic environment¹. Most macrophyte species are cosmopolitan, and groups of closely related species are known to replace each other in aquatic ecosystems around the world². These aquatic plants become weedy and grow profusely in water bodies, reducing their area and depleting water quality. For centuries, aquatic weeds were perceived as a nuisance rather than a resource because they block canals, hinder boat traffic, and increase waterborne diseases. In addition to serving as the base of aquatic food chain, these plants have many potential uses, such as human food, cosmetics, ethnomedicine, feeds for livestock, sources of fibre for paper making, and they are also used as fertilizer, like mulch and manure, ash, green manure, compost etc³.

Macrophytic vegetation includes deep-water and shallow-water species as well as species that actively grow in water in an emergent, submerged or floating state. These different categories have been reported to differ in chemical composition due to their relationship with the external environment. Aquatic plants commonly used for human consumption and livestock feed were evaluated by Ganai et al.⁴, Shah et al.⁵, and Rather and Nazir⁶, reporting high protein and micronutrient content in these weeds. Many researchers have explored the possibilities of using aquatic plants as a source of fodder for cattle^5,7, sheep^8,9, goats^10,11, poultry^12,13, and fish^14,15. The finding of these studies suggests that due to the acute shortage of fodder, these aquatic weeds can be used as a potential source of livestock feed.

In addition to being rich in protein, lipid, amino acids, and fatty acids, these macrophytes are an abundant source of minerals³. They serve as a good source of macro minerals like Calcium (Ca), Phosphorus (P), Sodium (Na), Magnesium (Mg), and microminerals like Iron (Fe), Manganese (Mn), Chromium (Cr), and Cobalt (Co)¹⁶. Minerals are essential catalysts for various biochemical reactions related to metabolism, growth, and development¹⁷. These mineral elements are separate entities from other essential nutrients like proteins, fats, carbohydrates, and vitamins. Each mineral has an optimum dose, and their concentration is critical to determining the achievement of the requirement in comparison with the health risks of deficiency and excess intake. Higher level of trace minerals required in minute quantities and even macrominerals like, Na, K, Mg, Ca, Fe, etc., may also be harmful¹⁷. Therefore, the chemical composition of macrophytes is essential to evaluate their food potential and nutritional value. On the other hand, aquatic plants accumulated heavy metals, improve water quality, and can be used as bio-indictors for trace element accumulation¹⁸. The present study aims to evaluate the mineral and heavy metal status of selected macrophytes from major water bodies namely Dal, Hokersar, Manasbal and Anchar lakes in Kashmir valley. This investigation will contribute to assessing the suitability of these macrophytes as feed ingredients in poultry and livestock nutrition.

Materials and methods

Study area

The study was conducted in the pristine aquatic ecosystems of Dal, Hokersar, Manasbal, and Anchar lakes, located in the picturesque region of Kashmir.

Dal lake: Situated in Srinagar (34.1106° N, 74.8683° E).

Characteristics: Spanning approximately 18 km², Dal Lake is an urban lake integral to tourism and recreation. It forms a crucial part of the natural wetland, contributing to the ecological diversity of the region.

Characteristics: A freshwater lake covering 2.81 km², Manasbal Lake boasts a maximum depth of 13 m (43ft) and a catchment area of 33 km². It presents a significant habitat for various aquatic species.

Hokersar lake: Positioned as a wetland conservation area (34°5′42″ N, 74°42′27″ E), 10 km northwest of Srinagar.

Characteristics: Covering an extensive area of 1375 hectares (13.75 km²), Hokersar plays a vital role in preserving the region’s wetland biodiversity.

Anchar lake: Situated in Srinagar district (34°9′0″N, 74°47′0″E).

Characteristics: This freshwater lake, with an area of 680 ha, faces ecological challenges, predominantly weed infestation. Despite its shallowness (maximum depth of 3 m), Anchar Lake serves as a unique ecosystem.

Collection and identification process

All the macrophytes used in this study were naturally growing in the water bodies of Kashmir without any human intervention. These aquatic plants occur profusely in lakes and waterways throughout the region. The water bodies are under the jurisdiction of the Lake Conservation and Management Authority (LCMA), Srinagar and collections were carried out only after obtaining prior permission from the Authority. The use of plant material in this study complied with the institutional guidelines of SKUAST-Kashmir, and all relevant national and international regulations.

The collection and identification of aquatic weeds for this study involved systematic procedures to ensure accuracy and reliability.

Site selection:

• Sites were carefully chosen in Dal, Hokersar, Manasbal, and Anchar lakes based on abundance and ease of macrophyte collection.

• In Dal Lake, samples were collected from four basins: Hazratbal, Nishat, Gagribal, and Nigeen basins.

• Other lakes under study also had four different selected sites for sampling.

• Multiple subsamples were collected from different locations within the same lake and pooled to form one composite sample to ensure representativeness and reduce within-site variability. Statistical replication was achieved by collecting independent composite samples from four lakes, namely Dal Lake, Manasbal Lake, Hokersar Lake, and Anchar Lake (n = 4).

Harvesting procedures:

• Collection of macrophytes followed the procedures developed at the Indian Institute of Science for aquatic ecosystems in India, including both streams and lakes.

• Different harvesting methods were employed based on the systemic forms of macrophytes.

• Emergent vegetation was harvested by clipping the above-ground standing crop.

• For submerged vegetation, a 1 m wooden pole was immersed in water where depth exceeded 1 m; in shallow areas, plants were manually collected.

• One representative sample of available macrophyte from each selected site was collected and were combined in one aggregate sample for each lake.

• Post-collection handling and management of samples were carried out in accordance with standard practices outlined under Aquatic Weed Management in India.

Transport and Preparation:

• After collection, samples were promptly transported to the laboratory.

• The collected weeds were washed thoroughly to remove adherent periphyton, detritus and marl, especially for rooted floating and submerged plants.

• A Teepol solution was used for washing, followed by rinsing with deionized water.

Drying and processing:

• The washed plants were then oven-dried at 60 °C for 72 h to ensure complete dehydration.

• Dried samples were ground to approximately 40 μm particle size for further analysis.

• Combustion in a muffle furnace at 480 °C was carried out, followed by mixing with a 32% HCl in a 50:50 solution.

Extraction:

• Extraction was performed using a Whatman filter paper, following the methodology outlined by Campbell and Plank¹⁹.

Identification:

• Taxonomic identification of collected macrophytes was carried out using appropriate keys, guides, and expert assistance.

• Species identification was carried out by Dr. Abdul Hamid Wani, Professor, Division of Taxonomy, University of Kashmir, Srinagar.

• A herbarium was prepared and deposited in the Division of Animal Nutrition, Faculty of Veterinary Sciences & Animal Husbandry, SKUAST-Kashmir, for future reference (Herbarium ID No. FVSc/ANN/Res/22–23/105–121).

• Each identified species was cataloged and associated with the specific sampling site.

This systematic approach ensures the reliability of data gathered and provides a comprehensive understanding of the mineral and heavy metal status of the selected macrophytes in the mentioned lakes.

Chemical composition of macrophytes

All the macrophytes after collection were analysed for proximate composition²⁰. The fiber fractions viz., neutral detergent fiber (NDF), acid detergent fiber (ADF), cellulose, hemicellulose and lignin, were evaluated as per the protocol proposed by Van Soest et al.²¹.

The calculation of total carbohydrates and non-fibrous carbohydrates was done by using formulae given by Sniffen et al.²², as under:

The chemical composition of macrophytes collected from different lakes under study is presented in Table 1.

Table 1.

Proximate composition and fiber fraction of major aquatic weeds from water bodies of central Kashmir.

Water body	Aquatic weeds	Proximate composition							Fiber fraction
		DM	CP	CF	EE	Ash	TC	NFC	NDF	ADF	Cellulose	HC	Lignin
DAL LAKE	Alternanthera philoxeroides	9.76 ± 1.28	13.66 ± 0.47	15.00 ± 0.23	3.20 ± 0.43	13.04 ± 0.13	70.00 ± 0.85	47.50 ± 1.54	22.51 ± 1.09	16.36 ± 0.52	13.87 ± 0.82	6.19 ± 1.05	11.95 ± 0.78
	Nymphaea tetragona	8.63 ± 0.88	7.07 ± 0.06	52.28 ± 3.24	3.90 ± 1.01	12.88 ± 2.11	75.20 ± 0.75	25.70 ± 0.15	49.51 ± 0.74	35.24 ± 0.40	14.48 ±  0.69	14.29 ± 0.92	12.86 ± 1.25
	Typha angustata	10.70 ± 1.02	10.07 ± 0.39	53.10 ± 1.67	1.93 ± 0.31	11.80 ± 0.50	75.90 ± 0.71	29.80 ± 0.95	46.15 ± 0.25	32.92 ± 0.52	14.84 ± 0.59	13.22 ± 0.27	12.32 ± 0.81
	Nymphoides peltata	8.53 ± 0.91	17.93 ± 0.21	36.73 ± 1.54	3.20 ± 0.26	13.13 ± 0.21	65.50 ± 0.33	21.20 ± 0.93	44.38 ± 0.80	32.52 ± 0.98	29.46 ± 1.10	11.86 ± 0.82	24.27 ± 0.88
	Nelumbo nucifera	9.80 ± 1.22	18.93 ± 0.37	8.53 ± 0.82	1.37 ± 0.31	11.70 ± 1.55	68.70 ± 0.64	56.30 ± 1.20	12.40 ± 0.99	7.75 ± 0.86	5.49 ± 0.69	4.64 ± 0.19	3.50 ± 0.55
	Myriophyllum spicatum	9.21 ± 0.81	17.20 ± 0.43	18.47 ± 0.82	1.83 ± 0.17	11.67 ± 0.40	69.10 ± 0.61	47.70 ± 1.15	21.35 ± 0.79	15.42 ± 0.92	12.42 ± 0.84	5.93 ± 0.14	9.39 ± 0.71
	Cladophora glomerata	8.92 ± 0.82	20.91 ± 0.82	18.90 ± 1.50	1.36 ± 0.41	15.42 ± 1.25	62.80 ± 1.15	39.50 ± 0.36	23.33 ± 0.79	17.33 ± 0.88	15.22 ± 0.73	6.01 ± 0.18	10.40 ± 0.62
	Salvinia natans	10.23 ± 0.97	18.07 ± 0.34	20.90 ± 1.65	2.30 ± 0.36	12.03 ± 0.21	67.50 ± 0.56	41.90 ± 1.67	25.61 ± 1.38	20.35 ± 0.76	18.28 ± 0.76	5.27 ± 0.66	12.41 ± 0.85
	Trapa natans	9.72 ± 0.92	4.11 ± 0.62	12.68 ± 1.75	0.70 ± 0.21	4.06 ± 0.78	91.10 ± 1.53	74.60 ± 2.37	16.52 ± 1.05	10.22 ± 0.75	7.21 ± 0.82	6.30 ± 0.30	5.44 ± 0.93
	Potamogeton lucens	10.74 ± 0.88	17.07 ± 0.34	23.27 ± 1.46	2.10 ± 0.26	11.77 ± 0.47	68.70 ± 0.57	39.60 ± 1.17	29.18 ± 0.78	23.16 ± 0.83	20.28 ± 0.94	6.01 ± 0.12	13.44 ± 0.97
	Nymphoides aquatica	9.34 ± 0.88	22.13 ± 1.63	23.02 ± 1.64	2.10 ± 0.16	19.92 ± 1.59	55.80 ± 2.97	26.50 ± 2.47	29.33 ± 0.87	21.19 ± 0.80	17.85 ± 1.26	8.14 ± 0.2	11.21 ± 0.78
	Ceratophyllum demersum	10.34 ± 0.79	14.27 ±0.34	24.80 ± 0.85	2.80 ± 0.22	12.57 ± 0.33	70.30 ± 0.87	41.90 ± 1.39	28.35 ± 0.74	20.42 ± 0.81	17.23 ± 0.77	8.98 ± 1.58	8.56 ± 1.31
	Potamogeton crispus	9.69 ± 0.86	12.70 ±0.65	21.59 ± 1.19	3.00 ± 0.58	21.70 ± 0.51	62.70 ± 0.90	37.50 ± 0.30	25.23 ± 0.78	17.59 ± 0.88	14.29 ± 0.68	7.64 ± 0.12	8.4 0.93
	Lemna minor	8.64 ± 0.91	11.32 ±1.27	14.05 ± 1.39	1.60 ± 0.30	15.18 ± 0.68	71.80 ± 1.96	50.50 ± 2.06	21.31 ± 0.88	14.28 ± 0.71	12.28 ± 0.88	7.03 ± 0.18	7.78 ± 0.81
MANASBAL LAKE	Potamogeton natans	8.51 ± 0.95	12.50 ± 0.98	12.24 ± 0.98	2.57 ± 0.72	19.30 ± 0.67	65.60 ± 2.34	41.10 ± 3.38	24.49 ± 1.05	15.17 ± 0.89	12.33 ± 1.02	9.32 ± 0.16	7.24 ± 0.78
	Nymphaea tetragona	8.26 ± 0.91	16.45 ± 1.05	15.41 ± 0.99	3.17 ± 0.72	12.49 ± 0.76	67.90 ± 2.51	43.60 ± 3.38	24.25 ± 0.83	15.24 ± 0.93	12.23 ± 0.78	9.01 ± 0.15	6.57 ± 0.84
	Nelumbo nucifera	9.36 ± 0.94	14.30 ± 0.36	14.23 ± 0.45	4.00 ± 0.54	10.16 ± 0.71	71.50 ± 1.53	57.20 ± 1.48	14.31 ± 1.07	7.16 ± 0.84	3.82 ± 0.69	7.15 ± 0.22	3.83 ± 0.85
	Potamogeton pectinatus	8.67 ± 0.96	11.47 ± 1.08	12.60 ± 0.90	2.93 ± 0.65	19.19 ± 0.87	66.40 ± 2.59	40.20 ± 3.49	26.18 ± 0.91	14.42 ± 0.95	11.29 ± 0.73	11.76 ± 0.20	5.42 ± 0.80
	Ceratophyllum demersum	8.59 ± 0.76	14.10 ± 0.36	12.73 ± 0.50	3.13 ± 0.58	12.31 ± 0.90	70.40 ± 1.76	48.10 ± 1.66	22.29 ± 0.74	15.27 ± 0.72	13.37 ± 0.66	7.03 ± 0.02	6.36 ± 0.99
	Myriophyllum spicatum	9.69 ± 1.04	14.03 ± 0.37	16.30 ± 0.54	2.13 ± 0.25	18.45 ± 0.74	65.30 ± 1.28	41.10 ± 1.18	24.26 ± 0.86	12.76 ± 0.78	9.22 ± 0.79	11.5 ± 0.08	3.54 ± 0.68
	Potamogeton natans	9.14 ± 0.82	13.93 ± 0.37	15.37 ± 0.56	2.50 ± 0.43	10.54 ± 0.78	73.00 ± 1.40	47.80 ± 1.17	25.16 ± 0.8	13.2 ± 0.78	10.21 ± 0.83	11.95 ± 0.019	4.29 ± 0.71
	Azolla cristata	8.54 ± 1.03	23.64 ± 1.16	13.55 ± 1.07	2.67 ± 0.63	17.31 ± 0.90	56.30 ± 2.67	15.90 ± 3.63	40.41 ± 0.98	27.38 ± 0.87	6.69 ± 0.69	13.03 ± 0.11	10.46 ± 1.04
HOKARSAR LAKE	Phragmites australis	9.58 ± 0.94	10.57 ± 0.99	26.24 ± 0.85	1.80 ± 0.68	15.43 ± 0.95	72.20 ± 2.60	34.90 ± 3.52	37.30 ± 0.92	25.36 ± 0.91	22.49 ± 0.99	11.94 ± 0.01	10.52 ± 1.09
	Azolla cristata	8.81 ± 0.7	22.20 ± 0.74	14.53 ± 0.93	3.36 ± 0.89	16.54 ± 0.90	57.90 ± 2.53	19.70 ± 3.32	38.17 ± 0.8	26.39 ± 1.38	5.37 ± 1.04	11.78 ± 1.44	12.66 ± 1.09
	Myriophyllum spicatum	8.59 ± 0.75	14.00 ± 0.41	15.27 ± 0.34	2.07 ± 0.37	17.05 ± 0.71	66.80 ± 1.48	41.60 ± 1.31	25.21 ± 0.79	16.21 ± 0.78	13.27 ±0.70	9.00 ± 0.01	6.40 ±1.08
	Crisium arvense	9.23 ± 0.87	11.68 ± 0.93	23.31 ± 0.87	1.77 ± 0.77	13.44 ± 0.73	73.10 ± 2.42	36.80 ± 3.35	36.27 ± 0.94	27.35 ± 0.94	24.34 ± 0.71	8.92 ± 0.06	8.82 ± 0.81
	Rumex rupestris	10.71 ± 0.94	23.53 ± 0.99	34.37 ± 1.05	2.20 ± 0.65	15.15 ± 0.83	59.10 ± 2.47	12.70 ± 3.37	46.33 ± 0.91	35.31 ± 0.96	32.32 ± 0.86	10.99 ± 0.06	9.60 ± 1.16
	Dysphania ambrosioides	11.07 ± 0.73	27.26 ± 1.03	23.40 ± 0.97	7.44 ± 0.02	17.29 ± 0.82	38.00 ± 2.85	2.45 ± 3.93	35.57 ± 1.09	26.41 ± 1.11	23.48 ± 1.12	9.16 ± 0.14	9.46 ± 0.77
ANCHAR LAKE	Azolla cristata	8.15 ± 1.41	20.85 ± 1.16	13.57 ± 1.26	3.22 ± 1.14	15.24 ± 1.21	60.60 ± 3.49	24.40 ± 5.11	36.20 ± 1.63	25.41 ± 1.28	4.53 ± 1.27	13.99 ± 1.31	10.85 ± 1.36
	Myriyophyllum spicatum	9.26 ± 1.33	12.23 ± 0.78	14.61 ± 1.30	2.30 ± 1.19	15.65 ± 1.20	69.80 ± 3.14	45.50 ± 4.68	24.27 ± 1.54	16.69 ± 1.11	13.36 ± 1.65	8.14 ± 1.27	7.29 ± 1.24
	Nymphaea tetragona	8.75 ± 1.59	7.01 ± 0.81	44.47 ± 1.77	3.21 ± 1.35	12.16 ± 1.35	77.60 ± 3.51	29.40 ± 4.99	48.20 ± 1.47	31.29 ± 1.19	14.21 ± 1.54	13.78 ± 1.28	12.53 ± 1.34
	Nymphoides peltata	8.67 ± 1.27	17.41 ± 0.88	35.26 ± 1.47	3.11 ± 1.47	13.25 ± 1.25	66.20 ± 3.57	23.80 ± 5.15	42.39 ± 1.58	27.44 ± 1.66	28.23 ± 1.77	11.61 ± 1.37	23.15 ± 1.62
	Nelumbo nucifera	9.48 ± 1.14	17.51 ± 1.01	8.31 ± 1.13	1.46 ± 1.29	10.44 ± 1.28	70.50 ± 3.52	58.10 ± 4.84	12.47 ± 1.33	8.20 ± 1.06	6.39 ± 1.20	6.18 ± 1.31	4.23 ± 1.37
	Salvinia natans	8.53 ± 1.54	18.29 ± 1.06	20.36 ± 1.18	2.37 ± 1.24	11.46 ± 1.76	67.80 ± 4.06	43.50 ± 5.43	24.32 ± 1.38	18.12 ± 1.36	17.21 ± 1.70	6.22 ± 1.23	12.21 ± 1.39
	Potamogeton natans	7.80 ± 1.73	16.76 ± 1.02	22.34 ± 1.20	2.45 ± 1.17	11.21 ± 1.38	69.50 ± 3.56	42.40 ± 4.96	27.16 ± 1.39	22.25 ± 1.34	19.22 ± 1.58	20.44 ± 1.59	13.45 ± 1.55
	Nymphoides aquatica	8.64 ± 1.16	19.67 ± 1.10	22.97 ± 1.60	1.78 ± 1.18	17.09 ± 1.32	61.40 ± 3.60	32.10 ± 4.78	29.36 ± 1.20	19.48 ± 1.10	18.66 ± 1.71	17.37 ± 1.32	10.54 ± 1.43
	Ceratophyllum demersum	9.26 ± 1.46	13.36 ± 1.28	20.26 ± 1.39	2.59 ± 1.51	12.34 ± 1.10	71.70 ± 3.88	45.10 ± 5.17	26.59 ± 1.29	16.24 ± 1.66	15.02 ± 1.37	18.23 ± 1.35	9.45 ± 1.17
	Lemna minor	8.19 ± 1.40	10.27 ± 1.17	12.52 ± 1.13	1.94 ± 1.51	15.07 ± 1.52	72.70 ± 4.19	52.40 ± 5.51	20.30 ± 1.33	12.51 ± 1.56	12.33 ± 1.32	12.54 ± 1.55	8.12 ± 1.44

Values are expressed as mean ± SEM of four replicates (n = 4).

DM. dry matter; CP. crude protein; CF. crude fiber; EE. ether extract; TC. total carbohydrates; NFC. non fiber carbohydrates NDF. neutral detergent fiber; ADF. acid detergent fiber; HC. hemicellulose.

Analysis of macro-minerals, micro-minerals, and heavy metals

Digestion process

The digestion of plant samples was conducted following the method outlined by Ahmed et al.⁸. A powdered plant sample (1 g) underwent digestion using a diacid mixture consisting of nine parts nitric acid to four parts perchloric acid. The digestion process occurred at approximately 80 °C until dense fumes appeared. Once digested, the samples were cooled to ambient temperature and transferred to a 50 ml volumetric flask. The volume was adjusted by adding Millipore ultrapure water to achieve proper dilution.

Analysis procedure

The digested samples were analyzed for their respective (Ca, P, Mg, Na, and K), micro-minerals (Cu, Zn, Fe, Co, and Mn), and heavy metals (Pb, Cr, As, and Hg) using an Atomic Absorption Spectrometer (AAS) (Agilent GTA 120, USA). The analysis was conducted in triplicate to ensure accuracy and reproducibility of the results.

In-vitro studies

Experimental animals

The animals for experiment were procured from sheep sub-unit of MRCS&G, Shuhama, SKUAST-K (34.1886°N; 74.8286°E), situated at an altitude of 5,212 ft above sea level. The experiment was approved by the Institutional Animal Ethics Committee (IAEC) of SKUAST Kashmir (1809/GO/ReL/15/CPCSEA) vide No AU/FVS/PS-57/3393. The animals were strictly handled as per regulations of committee for the purpose of control and supervision of Experiment on Animals (CPCSEA). Three adult male Corriedale sheep, aged between 18 and 24 months and exhibiting uniform conformation, were selected as donors of rumen inoculum for the in-vitro study. They were housed in a well-ventilated, hygienic environment with individual feeding arrangements. Prophylactic doses of Panacur^® (Fenbendazole) were administered as anthelmintic treatment. Periodic examinations of faecal and blood smears were conducted to monitor parasitic infestation. The animals were provided diets according to ICAR (2013)²³ guidelines to fulfil their nutrient requirements.

Sampling of rumen liquor

Rumen liquor was collected from the animals before feeding using a stomach tube, employing negative pressure generated by a suction pump²⁴. The collected rumen liquor individually underwent straining through four layers of muslin cloth [referred to as Strained Rumen Liquor (SRL)] and was then transported to the laboratory in a pre-warmed (39 °C) thermos flask. In-vitro studies were carried out in triplicate by transferring individual samples into pre-warmed (39 °C) fermentation flasks, following a modified method outlined by Tilley and Terry²⁵, and extraction was performed using a neutral detergent solution.

The incubation periods of 24, 48, and 72 h were selected to represent different phases of ruminal fermentation. The 24 h incubation reflects the extent of rapid and readily fermentable substrate degradation, whereas the 48 and 72 h incubations account for the progressive and near-complete degradation of slowly fermentable components.

Determination of in-vitro digestibility of aquatic weeds

Incubation of samples was conducted in 100 ml Erlenmeyer flasks. Finely ground samples (< 1 mm) of respective aquatic weeds (0.5 g) were taken in triplicate, along with 10 ml of SRL and 40 ml of McDougall buffer (composition: 9.80 g/l NaHCO3, 7.0 g/l Na2HPO4.2H2O, 0.47 g/l NaCl, 0.57 g/l KCl, 0.04 g/l CaCl2, and 0.12 g/l MgSO4.2H2O). Carbon dioxide was flushed through the contents of each vessel for 10 s, followed by immediate sealing with a stopper fitted with a control valve. The flasks were then incubated at 39 °C with periodic shaking for 48 h. After incubation, the contents of each flask were transferred into a spoutless beaker (1000 ml), washed thoroughly with neutral detergent solution, and the final volume in the beaker was adjusted to 150 ml. The contents were refluxed for 1 h at 100 °C, filtered through pre-weighed Gooch crucibles (G-I grade, 50 ml capacity), washed with distilled water, dried at 100 °C for 24 h, and weighed. Simultaneously, blanks were also incubated in triplicate alongside the standard. For in vitro studies at 72 h of incubation, samples were processed, and the reaction was stopped by adding 2 ml of 6 N hydrochloric acid and 0.1 g of pepsin powder (1:3000) to each vessel.

For determination of Dry Matter (DM), Organic Matter (OM), and Neutral Detergent Fibre (NDF) digestibility, Gooch crucibles containing undigested residue (NDF) were oven-dried at 100 °C for 24 h, cooled in desiccators, and weighed. The loss in dry matter and NDF was recorded as digested dry matter and digested NDF, respectively. The crucibles containing residue were then ignited in a muffle furnace at 600 °C, and the remaining quantity in the crucible after ignition (ash) was subtracted from the residual dry matter to determine the organic matter content.

Statistical analysis

The experimental data was evaluated by one-way ANOVA (SPSS Software, Base 23.0, for MacOS). The statistical equation used were as following:

where Yij is the parameter under analysis ij, µ is the overall mean, Ti is the effect due to treatments on the parameter under analysis, eij is the experimental error for ij on the observation. The significance of mean difference was tested by Duncan’s New Multiple Range Test (DNMRT). The significance of data was considered at p < 0.05. The data is presented in the table as mean, SEM and p-values.

Results

Macro-mineral composition

In the study encompassing fourteen freshwater macrophytes from Dal Lake (Table 2), Lemna minor exhibited significantly higher levels of Ca, P, Na, and K compared to other macrophytes. However, Trapa natans showed a significantly higher Mg level, followed by Potamogeton natans. Moving to Manasbal Lake (Table 3), Azolla cristata demonstrated significantly higher levels of Ca, P, Na, and K. For Mg content, Potamogeton natans and Potamogeton lucens exhibited the highest levels, followed by Potamogeton pectinatus and Ceratophyllum demersum. Among the six macrophytes studied in Hokersar Lake (Table 4), Rumex rupestris displayed significantly higher (p < 0.05) levels of Ca, while Azolla cristata had the highest P content. Phragmites australis showed the highest Na concentration, and Rumex rupestris displayed the highest Mg level. Azolla cristata had the significantly higher(p < 0.05) K content. In the case of Anchar Lake (Table 5), Lemna minor and Azolla cristata showed significantly higher (p < 0.05) levels of Ca, Na, and K. Azolla cristata, Nelumbo nucifera, Nymphoides aquatica, and Lemna minor had significantly higher (p < 0.05) P content. For Mg, Potamogeton lucens exhibited the highest levels, followed by Nymphoides aquatica, Ceratophyllum demersum, and Nymphoides peltata. The comparison of various macro-minerals present in common weeds from different lakes of Kashmir valley is presented in Fig. 1. These findings highlight the variability in macro-mineral composition among different macrophytes in each lake, shedding light on the potential nutritional diversity and suitability of these aquatic plants for various applications.

Table 2.

Macro-mineral, micro-mineral and heavy metal status of major aquatic weeds from Dal lake.

Attributes	Aquatic weeds														SEM	p-value
	AP	NT	TA	NP	NN	MS	CG	SN	TN	PN	NA	CD	PC	LM
Macro-mineral status (g/kg of dry weight)
Calcium	4.25ab	5.33abc	3.56ab	5.18abc	5.16abc	7.38c	5.68bc	5.49abc	4.30ab	5.00ab	3.20a	4.34ab	5.19abc	11.25d	0.33	< 0.05
Phosphorus	3.35abcd	2.24ab	2.24ab	3.54abcd	4.38bcd	2.51abc	4.83cd	3.37abcd	3.44abcd	3.71abcd	4.49bcd	1.73a	2.86abc	5.34d	0.22	0.028
Sodium	0.57a	2.46ab	0.98a	0.62a	2.17ab	1.45a	1.88ab	2.11ab	0.98a	0.61a	0.59a	3.63b	2.04ab	20.46c	0.78	< 0.05
Magnesium	1.63ab	1.13a	2.35abc	5.50fg	4.01cdef	3.13abcd	30.11i	2.91abcd	8.25h	7.15gh	5.13ef	4.85def	5.36fg	3.32bcde	1.09	< 0.05
Potassium	14.17cd	13.73bcd	11.78ab	12.03ab	12.73abcd	14.46de	11.82ab	11.11a	11.21a	11.42a	12.29abc	12.24abc	16.13e	24.67f	0.54	< 0.05
Micro-mineral status (ppm of dry weight)
Copper	8.48c	4.64b	8.04c	9.21c	9.07c	9.04c	8.92c	8.52c	8.18c	9.10c	0.39a	8.24c	9.10c	8.35c	0.38	< 0.05
Iron	211.14g	347.44l	115.14a	236.39k	199.09f	216.70h	210.13g	125.14b	140.13c	227.69j	158.45d	223.44i	161.50e	216.50h	8.83	< 0.05
Zinc	2.12a	8.48c	16.49f	11.49d	16.52f	16.53f	15.36ef	11.52d	16.25f	13.52e	5.47b	14.34ef	16.02f	15.44ef	0.70	< 0.05
Manganese	50.55a	132.25j	60.24b	122.34i	142.38k	90.21g	133.13j	62.33c	92.24h	90.38gh	122.26i	82.60e	70.23d	85.37f	4.52	< 0.05
Cobalt	2.43	1.84	0.71	1.14	1.64	0.30	2.14	1.17	2.27	2.24	1.29	1.13	1.76	2.20	0.16	0.376
Heavy metal status (ppm of dry weight)
Lead	0.48a	2.71bc	3.13bc	3.12bc	4.24cd	3.23bc	4.07bcd	3.18bc	3.91bcd	4.05bcd	2.19ab	0.43a	5.63d	3.17bc	0.24	< 0.05
Chromium	2.44ab	3.16b	0.49a	3.21b	3.11b	3.29b	3.15b	0.81a	2.15ab	2.34ab	3.17b	3.85b	3.44b	3.13b	0.20	0.029
Arsenic	86.64i	79.61h	56.01f	114.57k	42.38c	33.12b	95.69j	52.80e	42.50cd	59.23g	44.53d	30.52a	32.53ab	52.30e	3.88	< 0.05
Mercury	0.79	0.84	1.10	1.06	1.17	1.02	1.08	1.07	0.93	1.01	0.93	0.97	0.96	1.01	0.09	1.000

Values are expressed as mean and SEM of four replicates (n = 4).

Means with different superscripts between the columns differ significantly (p ≤ 0.05) as determined by one-way ANOVA followed by Duncan’s post-hoc test.

AP Alternanthera philoxeroides, NT Nymphaea tetragona, TA Typha angustata, NP Nymphoides peltata, NN Nelumbo nucifera, MS Myriophyllum spicatum, CG Cladophora glomerata, SN Salvinia natans, TN Trapa natans, PN Potamogeton natans, NA Nymphoides aquatica, CD Ceratophyllum demersum, PC Potamogeton crispus, LM Lemna minor.

Table 3.

Macro-mineral, micro-mineral and heavy metal status of major aquatic weeds from Manasbal lake.

Attributes	Aquatic weeds								SEM	p-value
	PN	NT	NN	PP	CD	MS	PL	AC
Macro-mineral status (g/kg of dry weight)
Calcium	4.23bc	3.11ab	2.89ab	4.91c	2.91ab	4.44bc	2.43a	9.74d	0.48	< 0.05
Phosphorus	2.22abc	1.68ab	3.70cd	2.21abc	1.16a	2.15abc	3.18bc	4.86d	0.27	0.003
Sodium	1.30a	1.90ab	1.76ab	1.86ab	3.19b	0.98a	0.42a	18.35c	1.17	< 0.05
Magnesium	5.38d	0.92a	3.87bcd	4.88cd	4.24bcd	2.80b	5.48d	3.15bc	0.33	< 0.05
Potassium	15.22c	9.67ab	8.78a	14.37c	9.61ab	10.67b	8.50a	22.36d	0.93	< 0.05
Micro-mineral status (ppm of dry weight)
Copper	7.47bc	3.23a	7.19bc	8.18c	6.22b	8.20c	8.59c	8.06c	0.37	< 0.05
Iron	158.18a	344.20h	193.20c	161.30b	221.20f	208.31d	223.28g	213.16e	11.28	< 0.05
Zinc	13.25bcd	7.17a	15.17e	14.22cde	13.06bc	15.10de	12.21b	14.17cde	0.53	< 0.05
Manganese	67.29a	129.39e	140.18f	66.90a	78.23b	88.18d	88.52d	83.17c	5.34	< 0.05
Cobalt	1.25	1.35	1.24	1.22	1.25	0.37	2.04	2.05	0.15	0.119
Heavy metal status (ppm of dry weight)
Lead	3.09a	4.28a	3.08a	3.49a	4.56a	6.41b	4.20a	3.87a	0.25	0.003
Chromium	3.52c	2.17bc	3.07c	2.52c	3.02c	2.87c	0.45a	0.90ab	0.25	0.003
Arsenic	39.25e	37.70de	42.44f	36.37cd	30.47a	33.06b	32.50b	35.79c	0.77	< 0.05
Mercury	0.83	0.82	0.88	0.89	0.81	0.84	0.91	0.88	0.06	1.000

Values are expressed as mean and SEM of four replicates (n = 4).

Means with different superscripts between the columns differ significantly (p ≤ 0.05) as determined by one-way ANOVA followed by Duncan’s post-hoc test.

PN, Potamogeton natans; NT, Nymphaea tetragona; NN, Nelumbo nucifera; PP, Potamogeton pectinatus; CD, Ceratophyllum demersum; MS, Myriophyllum spicatum; PL, Potamogeton lucens; AC, Azolla cristata.

Table 4.

Macro-mineral; micro-mineral and heavy metal status of major aquatic weeds from Hokersar lake.

Attributes	Aquatic weeds						SEM	p-value
	PA	AC	MS	CA	RR	DA
Macro-mineral status (g/kg of dry weight)
Calcium	2.84a	9.43c	5.17b	4.70ab	10.11c	3.44ab	0.71	< 0.05
Phosphorus	3.25a	4.87b	2.02a	2.03a	3.54ab	2.69a	0.29	0.012
Sodium	20.32d	17.65c	0.98a	1.62ab	2.70b	2.28ab	1.97	< 0.05
Magnesium	2.85a	3.11a	2.33a	3.32a	4.81b	3.46ab	0.24	0.043
Potassium	14.35d	22.26f	10.72c	16.61e	5.40a	8.43b	1.35	< 0.05
Micro-mineral status (ppm of dry weight)
Copper	8.63b	7.63b	8.04b	7.51b	2.28a	7.88b	0.54	< 0.05
Iron	210.22c	212.25d	206.40b	205.22b	40.05a	214.28e	15.35	< 0.05
Zinc	14.79c	13.71bc	14.42bc	12.35b	5.41a	13.29bc	0.80	< 0.05
Manganese	80.44c	82.39d	87.45f	71.15b	11.11a	85.47e	6.47	< 0.05
Cobalt	1.19	1.92	0.36	1.13	1.25	1.13	0.17	0.246
Heavy metal status (ppm of dry weight)
Lead	3.61	3.45	3.61	5.67	2.63	4.13	0.30	0.072
Chromium	3.15	3.36	4.28	3.75	4.17	3.28	0.21	0.603
Arsenic	48.77b	53.73c	43.10a	56.34d	62.48e	52.29c	1.47	< 0.05
Mercury	1.05	1.03	1.13	0.94	0.92	0.85	0.09	0.971

Values are expressed as mean and SEM of four replicates (n = 4).

Means with different superscripts between the columns differ significantly (p ≤ 0.05) as determined by one-way ANOVA followed by Duncan’s post-hoc test.

PA, Phragmites australis; AC, Azolla cristata; MS, Myriophyllum spicatum; CA, Crisium arvense; RR, Rumex rupestris; DA, Dysphania ambrosioides.

Table 5.

Macro-mineral, micro-mineral and heavy metal status of major aquatic weeds from Anchar lake.

Attributes	Aquatic weeds										SEM	p-value
	AC	MS	NT	NP	NN	SN	PL	NA	CD	LM
Macro-mineral status (g/kg of dry weight)
Calcium	9.33b	4.71a	5.32a	5.20a	5.06a	5.26a	4.64a	3.30a	3.78a	10.95b	0.47	< 0.05
Phosphorus	4.46	1.83	2.13	3.22	4.33	3.35	3.42	4.57	1.78	4.90	0.29	0.084
Sodium	15.20b	0.92a	2.37a	1.00a	2.27a	2.02a	0.92a	0.88a	3.28a	17.31b	1.11	< 0.05
Magnesium	2.57ab	2.26ab	1.18a	4.39bc	4.29bc	2.88ab	6.32c	4.70bc	4.75bc	3.09ab	0.36	0.034
Potassium	20.20b	10.18a	11.64a	10.95a	11.35a	10.56a	10.50a	11.31a	9.45a	19.25b	0.72	< 0.05
Micro-mineral status (ppm of dry weight)
Copper	7.26c	7.43c	4.45b	9.16c	8.62c	8.50c	8.60c	0.63a	7.54c	8.01c	0.50	< 0.05
Iron	207.52e	202.46d	343.39i	229.56h	191.68c	122.99a	228.98h	157.93b	222.37g	216.63f	10.06	< 0.05
Zinc	12.80cd	14.25cde	8.47b	10.96bc	16.56e	11.31bc	12.56cd	5.08a	13.37cde	15.25de	0.65	< 0.05
Manganese	81.44b	86.57cd	132.31f	121.29e	141.53g	60.59a	88.80d	120.70e	81.48b	83.49bc	4.73	< 0.05
Cobalt	1.92	0.69	1.95	1.48	1.52	1.67	2.36	1.71	1.49	1.92	0.27	0.992
Heavy metal status (ppm of dry weight)
Lead	3.37	3.47	2.58	3.25	4.33	2.90	3.69	2.35	0.68	3.20	0.27	0.198
Chromium	3.25	4.04	3.12	3.11	2.85	0.82	2.80	3.17	3.76	3.12	0.24	0.307
Arsenic	51.55d	41.46b	78.51f	112.46g	42.27b	56.17e	57.26e	45.88c	31.31a	51.47d	4.09	< 0.05
Mercury	1.07	1.34	1.13	1.15	1.57	1.27	1.12	1.39	1.36	1.34	0.21	1.000

Values are expressed as mean and SEM of four replicates (n = 4).

Means with different superscripts between the columns differ significantly (p ≤ 0.05) as determined by one-way ANOVA followed by Duncan’s post-hoc test.

AC, Azolla cristata; MS, Myriophyllum spicatum; NT, Nymphaea tetragona; NP, Nymphoides peltata; NN, Nelumbo nucifera; SN, Salvinia natans; PL, Potamogeton lucens; NA, Nymphoides aquatica; CD, Ceratophyllum demersum; LM, Lemna minor.

Fig. 1.

Comparison of macro-minerals among common macrophytes from different lakes of central Kashmir.

Micro-mineral composition

Among different aquatic macrophytes collected from Dal Lake the micro mineral concentration was almost similar with common pattern (Table 2). Cu concentration was lower in Nymphoides aquatica followed by Nymphaea tetragona with almost similar concentration among other weeds from Dal Lake. Among different microminerals, Fe was found in highest concentration in these collected macrophytes. Significantly highest (p < 0.05) concentration of Fe was reported in Nymphaea tetragona followed by Nymphoides peltata, Potamogeton lucens, Ceratophyllum demersum, Myriophyllum spicatum and Lemna minor. Zn concentration was found significantly lower in Alternanthera philoxeroides followed by Nymphoides aquatica, Nymphaea tetragona and Nymphoides peltata. The concentration of Zn in other macrophytes of Dal Lake was not significantly different (p≥0.05). The concentration of Mn also differed significantly (p < 0.05) among different macrophytes with highest concentration in Nelumbo nucifera followed by Cladophora glomerata, Nymphaea tetragona, Nymphoides peltata. Significantly lower Mn concentration was reported in Alternanthera philoxeroides and Typha angustata. Co concentration was similar among different macrophytes from Dal lakes without significant difference.

Among different macrophytes collected from Manasbal lake (Table 3), significantly lower (p < 0.05) level of Cu and higher level of Fe was reported in Potamogeton natans, whereas Zn and Mn content was significantly higher (p < 0.05) in Nelumbo nucifera with non-significant difference (p≥0.05) in Co concentration.

In macrophytes of Hokersar lake (Table 4) significantly lower concentration of Cu was reported in Rumex rupestris with non-significant difference among other five macrophytes. Dysphania ambrosioides had significantly higher (p < 0.05) concentration of Fe followed by Azolla cristata, Phragmites australis and Crisium arvense with lowest concentration in Rumex rupestris. Zn and Mn concentration was also found significantly lower (p < 0.05) in Rumex rupestris and highest Phragmites australis with non-significant difference (p≥0.05) in Co concentration among different macrophytes.

Cu concentration was significantly lower (p < 0.05) in Nymphoides aquatica, followed by Nymphaea tetragona in macrophytes collected from Anchar lake (Table 5). Significantly highest (p < 0.05) concentration of Fe was reported in Nymphaea tetragona followed by Nymphoides peltata, Potamogeton lucens, Ceratophyllum demersum, Lemna minor, Azolla cristata and Myriophyllum spicatum. Significantly highest (p < 0.05) Zn concentration was reported in Myriophyllum spicatum, with comparatively lower concentration in Nymphoides aquatica followed by Nymphaea tetragona, Nymphoides peltata and Salvinia natans. The concentration of Mn also differed significantly (p < 0.05) among different macrophytes with highest concentration in Nelumbo nucifera followed by Nymphaea tetragona, Cladophora glomerata, Nymphoides peltata and Nymphaea tetragona. Significantly lower Mn concentration was reported in Salvinia natans and Azolla cristata. Among microminerals Co was present in lowest concentration without significant difference (p≥0.05) among different macrophytes. The comparison of various micro-minerals present in common weeds from different lakes of Kashmir valley is depicted in Fig. 2.

Fig. 2.

Comparison of micro-minerals among common macrophytes from different lakes of central Kashmir.

Heavy metal status

The heavy metal status among macrophytes in various lakes reveals distinct concentrations of different elements. In Dal Lake, Potamogeton crispus stands out with a significantly higher (p < 0.05) concentration of Pb (Fig. 3), followed by Nelumbo nucifera, Cladophora glomerata, and Potamogeton lucens (Table 2). Conversely, exhibit significantly lower levels of Cr (Fig. 3). Nymphoides peltata reports the highest concentration of Ar (Fig. 3), while Ceratophyllum demersum shows the lowest. Surprisingly, the concentration of Hg does not vary significantly (p≥0.05) among these macrophytes (Fig. 3). In Manasbal Lake, Myriophyllum spicatum displays a significantly higher (p < 0.05) concentration of Pb, while Potamogeton lucens and Azolla cristata exhibit lower levels of Cr compared to other macrophytes (Table 3). The concentration of As differs significantly (p < 0.05), with Nelumbo nucifera having the highest, followed by Potamogeton natans, Nymphaea tetragona, Potamogeton pectinatus, and Azolla cristata. No significant difference (p≥0.05) was found in the concentration of Hg among the macrophytes.

Fig. 3.

Comparison of heavy metals among common macrophytes from different lakes of central Kashmir.

Hokersar water body and Anchar water body reveal non-significant differences (p≥0.05) in the levels of Pb, Cr, and Hg among the respective macrophytes. In Hokersar, Rumex rupestris exhibits significantly higher (p < 0.05) levels of As, followed by Crisium arvense, Azolla cristata, Dysphania ambrosioides, and Phragmites australis, while Myriophyllum spicatum displays lower levels. In Achar lake, Nymphoides peltata shows the highest concentration of As, followed by Nymphaea tetragona, Salvinia natans, Potamogeton lucens, Lemna minor, and Azolla cristata. Ceratophyllum demersum is noted for having lowest level of As in Anchar Lake.

In-vitro digestibility of aquatic weeds

The results of in-vitro digestibility of various aquatic weeds from different water bodies in Kashmir valley is presented in Table 6. Nelumbo nucifera, Trapa natans, and Lemna minor, sourced from Dal Lake, exhibited notably higher in-vitro digestibility percentages for dry matter (64.31%, 63.46%, and 60.23% respectively), organic matter (79.20%, 78.30%, and 75.40%), and NDF (69.19%, 68.22%, and 65.40%). Nymphaea tetragona, sourced from Manasbal Lake, also showed relatively high values (63.25%, 68.58%, and 58.47%). In contrast, Typha angustata, Nymphoides aquatica, and Ceratophyllum demersum, from Nigeen Lake, demonstrated lower in-vitro digestibility percentages: 35.40%, 40.39%, and 30.45% for dry matter; 40.38%, 45.28%, and 35.31% for organic matter; and 40.83%, 45.64%, and 35.54% for NDF, respectively. Similarly, Nymphoides peltata, sourced from Anchar Lake, exhibited lower values as well, with percentages of 42.30%, 47.25%, and 37.50%.

Table 6.

Percent in-vitro digestibility of major aquatic weeds from water bodies of central Kashmir.

Water body	Aquatic weeds	IVDMD	IVOMD	IVNDFD
DAL LAKE	Alternanthera philoxeroides	55.31e	70.31f	60.37f
	Nymphaea tetragona	54.25de	69.28ef	59.40ef
	Typha angustata	53.32cde	68.33def	58.32def
	Nymphoides peltata	46.51a	61.50a	51.52a
	Nelumbo nucifera	64.31g	79.22i	69.19h
	Myriophyllum spicatum	51.28bc	66.43bcd	56.45bcd
	Cladophora glomerata	50.37b	65.27b	55.30b
	Salvinia natans	52.25bcd	67.41cde	57.26bcd
	Trapa natans	63.46g	78.40i	68.22h
	Potamogeton lucens	50.31b	65.39bc	55.46bc
	Nymphoides aquatica	52.35bcd	67.42cde	57.53cde
	Ceratophyllum demersum	50.70b	65.53bc	55.99bc
	Potamogeton crispus	58.38f	73.44g	63.49g
	Lemna minor	60.23f	75.44h	65.40g
SEM		0.81	0.80	0.79
P value		< 0.05	< 0.05	< 0.05
MANASBAL LAKE	Potamogeton natans	61.41c	66.45cd	56.38c
	Nymphaea tetragona	63.25cd	68.58de	58.47cd
	Nelumbo nucifera	45.34a	50.45a	40.46a
	Potamogeton pectinatus	64.40d	69.43e	59.46d
	Ceratophyllum demersum	61.11c	66.29c	56.21c
	Myriophyllum spicatum	65.52d	70.39e	60.48d
	Potamogeton lucens	64.73d	69.58e	59.60d
	Azolla cristata	55.47b	60.59b	50.50b
SEM		1.34	1.33	1.33
P value		< 0.05	< 0.05	< 0.05
HOKERSAR LAKE	Phragmites australis	55.21b	60.46b	50.52b
	Azolla cristata	52.66a	57.47a	47.48a
	Myriophyllum spicatum	61.14c	66.30c	56.22c
	Crisium arvense	60.20c	65.32c	55.42c
	Rumex rupestris	54.54ab	59.45ab	49.17ab
	Dysphania ambrosioides	55.20b	60.25b	50.38b
SEM		0.78	0.81	0.82
P value		< 0.05	< 0.05	< 0.05
ANCHAR LAKE	Azolla cristata	49.25cd	54.82cd	44.14cd
	Myriyophyllum spicatum	60.37f	65.41f	55.08f
	Nymphaea tetragona	51.57d	56.36d	46.39d
	Nymphoides peltata	42.30a	47.25a	37.50a
	Nelumbo nucifera	60.76f	65.76f	55.76f
	Salvinia natans	47.15bc	52.09bc	42.33bc
	Potamogeton lucens	48.77cd	53.42bcd	43.51bcd
	Nymphoides aquatica	47.45bc	52.07bc	42.30bc
	Ceratophyllum demersum	45.11ab	50.24ab	40.36ab
	Lemna minor	56.15e	61.73e	51.32e
SEM		1.14	1.16	1.12
P value		< 0.05	< 0.05	< 0.05

Values are expressed as mean and SEM of four replicates (n = 4).

Means with different superscripts within rows differ significantly (p ≤ 0.05) as determined by one-way ANOVA followed by Duncan’s post-hoc test.

Discussion

Mineral content of these aquatic weeds collected from various water bodies, revealing diverse concentrations influenced by factors such as location and growth stages. The study indicates significant mineral content across all species, consistent with previous research of Khan et al.²⁶. Saraf²⁷ in his study about nutritional status of some common aquatic weeds in Dal and Nilnag lakes of Kashmir and reported comparable Ca concentration. Similar mineral composition was reported by Adelakun et al.²⁸. who reported that K concentration was the highest of all macro-mineral elements, though varied among the sampled plants of Nymphea lotus, Pistias tratioties, Eichorrnia crassipes, and Ipomoea aquatica. The study conducted by Kumar et al.¹⁶ investigated the mineral concentrations in various aquatic plants and reported that P. stratiotes exhibited the highest levels of sodium, magnesium, chromium, and iron. H. verticillata was identified as a rich source of copper, manganese, cobalt, and zinc. S. polyrhiza had the highest contents of calcium, magnesium, strontium, and nickel.

Macro-minerals

Among macro-minerals, potassium (K) and iron (Fe) emerge as the most abundant elements in these plants. Calcium (Ca) concentrations vary, with higher levels observed in Lemna minor from Dal and Anchar lakes²⁶, Rumex rupestris from Hokersar wetland, and Azolla cristata from Manasbal and Anchar lakes. Phosphorus (P) levels are highest in Lemna minor and lowest in Ceratophyllum demersum. Sodium (Na) concentration is elevated in Lemna minor from Dal and Nigeen lakes, and in Phragmites australis from Hokersar wetland. Azolla cristata from Hokarser wetland, Manasbal, and Anchar lakes also exhibits high Na concentration, while Potamogeton lucens has the lowest Na concentration. Cladophora glomerata from Dal and Nigeen lakes records the highest magnesium (Mg) content, whereas Nymphaea tetragona from Manasbal lake has the lowest. Lemna minor from Dal and Nigeen lakes and Azolla cristata from Manasbal and Anchar lakes exhibit high potassium (K) concentrations, whereas Potamogeton lucens and Nelumbo nucifera from Manasbal lake show the lowest K concentrations.

Micro-minerals

Microminerals show variation, with copper (Cu) concentrations being similar among many weeds, highest in Nymphoides peltata, and lowest in Nymphoides aquatica. Iron (Fe) concentration is highest in Nymphaea tetragona and lowest in Rumex rupestris from Hokersar wetland. Zinc (Zn) concentrations vary, with the highest in Nelumbo nucifera and Myriophyllum spicatum, and the lowest in Alternanthera philoxeroides. Factors such as soil, climate, species and stage of maturity contribute to variations in the concentration of minerals in forages^29,30. This remark corresponds well with our findings. Jabin et al.³¹. evaluated the micro-mineral (Cu, Co, Cr, Fe, Mn, Mo and Zn) concentration of six selected aquatic plants, namely Eichhornia crassipes, Hydrilla verticillata, Ipomoea aquatica, Nymphaea rubra, Pistia stratiotes, and Trapa natans and reported the concentrations of individual elements varied. These findings provide valuable insights into the mineral composition of different aquatic plant species, which can have implications for various applications, including environmental monitoring and nutrient management. It emphasizes the variability in mineral content across different plant species, highlighting their diverse roles in nutrient cycling and potential use in various applications such as phytoremediation or as nutrient-rich feed sources.

Heavy metals

The presence of essential elements in aquatic plants indicates their potential as nutrient sources. However, the study also highlights the potential accumulation of heavy metals such as lead (Pb), mercury (Hg), arsenic (As), and chromium (Cr), possibly originating from sewage discharges. Some plants, like Myriophyllum spicatum, show promise as phytoremediators for moderate Pb pollution. Similarly, Etse et al.³². determined the nutritional composition of Nymphaea lotus, Typha australis, Ipomoea aquatica and Scirpus cubensis, reported that heavy metals in the sediment samples revealed significant variations. Aquatic plants are recognized as pollution monitoring organisms due to their ability to accumulate heavy metals through their roots and shoots³³. Heavy metals are recognized as toxic pollutants entering aquatic environments, with the World Health Organization (WHO) recommending a limit of 2 mg/kg for Pb in aquatic plants, a limit exceeded by many plants in this study. High Pb concentrations were reported by Flefel et al.³⁴. in various aquatic plants, and similar observations were made in the plants of Taihu Lake by Bai et al.¹⁸. Dixit and Tiwari³⁵ reported comparable heavy metal accumulation in Water hyacinth. In a study conducted in Pariyej Community Reserve, Gujarat, Kumar et al.³⁶. reported varying concentrations of Pb in different macrophyte species, emphasizing the need for monitoring and addressing heavy metal pollution in diverse aquatic ecosystems.

In-vitro digestibility

The elevated in-vitro digestibility observed in these aquatic plants compared to land-based fodders can be attributed to their lower crude fiber and lignin content. In a study by Huque et al.³⁷, the digestibility of Spirodela, Lemna, and Wolffia, the most common duckweed species, was assessed. They found that the dry matter (DM) digestibility for 24 h was 41.00%, 57.00%, and 73.10% respectively, and the crude protein (CP) digestibility was 52.80%, 74.00%, and 77.80% DM respectively. Similarly, Khan et al.²⁶. investigated the nutritive potential of Lemna trisulaca, Lemna perpusila, Azolla pinnata, and Eichhornia crassipes. Their findings revealed that the organic matter digestibility after 24 h of incubation ranged between 529 and 556 g/kg DM among the species studied, with Azolla exhibiting the highest digestibility in this regard.

It is crucial to consider the impact of harvesting aquatic plants on mineral nutrient removal from aquatic environments. Additionally, the higher concentrations of heavy metals in aquatic weeds compared to terrestrial fodders underscore the need for environmental monitoring and management. The study aligns with existing literature on mineral concentrations in aquatic plants, emphasizing the importance of factors like soil, climate, species, and maturity stage in influencing mineral content.

Conclusion

In conclusion, among macro-minerals, potassium (K) and among micro-minerals iron (Fe) emerge as the most abundant elements in these plants. Ca, P, Na and K concentrations were higher in Lemna minor. Azolla cristata exhibits high Na concentration, while Cladophora glomerata has highest Mg content. In microminerals Cu concentrations being highest in Nymphoides peltata, and Fe concentration were highest in Nymphaea tetragona. While, Zn were found highest in Nelumbo nucifera and Myriophyllum spicatum. Myriophyllum spicatum, show promise as phytoremediators for moderate Pb pollution. Overall, the study provides valuable information about the ecological significance and potential applications of these macrophytes in various fields. Since some aquatic weeds with higher nutrient content (Dysphania ambrosoides, Trapa natans, Lemna minor, Nelumbo nucifera, Azolla spp.) comparable to leguminous fodders, with better biomass and digestibility can be effectively used as replacement of quality production fodders in animal diets. Further research and monitoring are essential to explore the dynamic interactions between aquatic plants and their environment, particularly in the context of changing environmental conditions and human activities.

Author contributions

M Islam: Investigation, Methodology, Visualisation. GG Sheikh: Concept, Visualisation, Investigation, Formal analysis. QS Sahib: Writing- original draft, Methodology, Supervision. HA Ahmed: Supervision, Validation.

Funding

Not applicable.

Data availability

All the data generated and analysed during this study are included in this manuscript.

Declarations

Competing interests

The authors declare no competing interests.

Ethics approval

The experiment was approved by the Institutional Animal Ethics Committee (IAEC) of SKUAST-Kashmir (1809/GO/ReL/15/CPCSEA) vide No AU/FVS/PS-57/3393. The present study was conducted and reported in accordance with the ARRIVE guidelines.