A NOVEL FERMENTED FOOD

THE YELLOW WATER LILY (NUPHAR ADVENA)

BY

SANDRA STURTON

A report submitted in partial fulfillment of the requirements of the Senior Project Course 336-4900

DEPT. AGR. ENG.

MACDONALD COLLEGE

MCGILL UNIVERSITY

MARCH 1979 ACKNOWLEDGEMENTS

I would like to thank Prof. Kok for his extensive assistance and guidance in this project.

I would also like to thank Profs. Smyrl, Beveridge, and Farmer for their assistance and advice in various aspects of this project.

I would especially like to thank Prof. Blackwood for his aid in the microbiological side of my project (the fermentation).

Finally, I want to extend many thanks to those who helped me harvest the roots, and to those who were panelists on my taste panel, particularly Teri who recruited several of the panelists for me.

- i - TABLE OF CONTENTS

PAGE

ACKNOWLEDGEMENTS i

TABLE OF CONTENTS i i

OBJECTIVES 1

LITERARY REVI ElAJ 2

WILD EDIBLE FOODS 3

YELLOW WATER LILY (NUPHAR ADVENA) 7 AQUATIC PLANTS - PESTS 9

FERMENTATION 11

EXPERIMENTAL WORK 15

COLLECTION AND TREATMENT 16 FERt·1ENTATI ON 17 MOISTURE CONTENT 25

ASH CONTENT 27

PROTEIN CONTENT 29

FAT CONTENT 31

MICE TRIAL 33 TASTE PANEL 35

DISCUSSION AND CONCLUSIONS 39 BIBLIOGRAPHY 41

- i i - OBJECTIVES

Many of the vegetables grown in North America are not indigenous to this area. Consequently, these plants must attempt to adapt to our conditions, and they must compete with weeds which grow native and thrive here. If we chose to grow indigenous plants, we could utilize submarginal land without constantly battling weeds and pests, stronger and more suited to the environment.

Water lilies are indigenous to this area and grow in conditions not suited to any of our other food crops. Often, they are a pest in themselves, in that they must be removed without being utilized. By utilizing water lilies as a food crop, we could generate food yields from marginal swamp and pond areas.

Water lily roots have been eaten in the past by Indians and health food enthusiasts; however, presently they are not commonly utilized. In this project, it has been attempted to create a novel food of good nutritive value and taste by fermenting them. Analyses of content were done before and after fermentation, and a taste panel was conducted.

- 1 - LITERARY REV! EvJ

- 2 - WILD EDIBLE FOODS

Most of the vegetables eaten today by Canadians are not indigenous to this area, having originated in considerably different climates and adapted as . best they could, to our climate.

The temperate parts of America had few native vegetables which were cultivated. Prehistoric indians raised corn, cassava, sweet

potatoes, pumpkins, and beans (Pederson, 1971). However~ America had to wait for foreign travellers before she met and grew many of the common vegetables we know today. Among the immigrants are peas {Central Europe), tomatoes (Peru), asparagus (Great Britain, Russia, Poland), be.ets (North Africa, West Asia), brocolli, brussel sprouts, cabbage (England, West European Coast), and carrots (Europe, West Asia). (Bianchini & Corbetta, 1976; Nissley, 1943) Additional plants and their origins can be found in Table 1.

When given these new crops, we accepted and made them our own, working long and hard to make them thrive in their new environment. Although these vegetables are successful in growing here, they must be

continuously cultivated and nurtured against the native, competitive~ dominant plants which need no encouragement to thrive here.

It would seem practical to study some of these indigenous wild plants as future food crops which could possibly produce greater yields without having to fight the competition as introduced vegetables must. Plants which could be grown on some of the marginal land here which introduced plants won't accept; but which native plants call home. - 3 - TABLE 1

SO ~ E COMMON VEGETABLES* _.:. ORIGINS AND CONTENTS Content % Venetables Origin Hater Protein Fat Ash Carbohydrates Jerusalem Artichoke N.or S. America 79.5 2.2 0.1 1 .17 17.0 Asparagus G. B., Russia, Poland 93.0 2.2 0.2 0.67 3.9 Beans America 88.9 2.4 0.2 0.77 7.7 Beets N.Africa, W.Asia 87.6 1 . 6 0.1 1 .11 9.6 Brocol1i W.European Coast 89.9 3.3 0.2 1 . 1 5.5 Brussel Sprouts W.European Coast 84.9 4.4 0.5 1. 28 8.9 Cabbage W.European Coast 92.4 1.4 0.2 0.75 5.3 Carrots Europe 88.2 1. 2 0.3 1. 02 9.3 Cassava Equatorial Amer. 11 . 85. Cauliflower Europe, H. Asia 91.7 2.4 0.2 0.85 4.9 Celery Canary Islands 93.7 1 .3 0.2 1.08 3.7 Swiss Chard H.Asia, Mediterr. 95.2 1. 0 0.1 0.8 2.9 Cucumbers E.Indies, India 96.1 0.7 0.1 0.44 2.7 Dandelion Temperate Countries 85.8 2.7 0.7 2.0 8.8 Eggplant S. America 92.7 1 . 1 0.2 0.54 5.5 Lettuce Asia 94.8 1 .2 0.2 0.91 2.9 Mushrooms 91 . 1 1.14 0.3 Muskmelons s. Asia 92.7 0.6 0.2 0.6 5.9

Onions ~~. Asia 87.5 1. 4 0.2 0.58 10.3 · Parsley Mediterranean 83.9 3.7 1. 0 2.4 9.0 Peanuts America 15. 25. 50. 2-3 Peas Central Eur. & Asia 74.3 6.7 0.4 0.92 17.7 - 4 - TABLE 1 (continued)

SO ME COMMON VEGETABLES* -- ORIGINS AND CONTENTS Content Vegetable Origin Water Protein Fat Ash Carbohydrates Pumpkin Tropi ea 1 America 90.5 1 .2 0.2 0.82 7.3 Spinach Persia 92.7 2.3 0.3 1 . 5 3.2 Squash Tropical America 88.6 1 . 5 0.3 0.83 8.8 Sweet Corn Peru 73.9 3.7 1 .2 0.66 20.5 Sweet Potatoes Tropi ea 1 America 68.5 1. 8 0.7 1. 07 . 27.9 Tomato Peru 94.1 1.0 0.3 0.57 4.0 Turnip 90.0 1 . 1 0.2 0. 73 7. 1

~Jh i te Potatoes S.America, Mexico 77.8 2.0 0. l 0.99 19.1

- 4 A - Only 1% of the world's edible plants are be{ng used routinely as food. Approximately 95 % of our domestic food comes from only

20 crops; in many c~untries, fewer than 6 crops are actually exploited. (Parrish, et al, 1974) Although there are numerous possibilities for different foods, many people are too set in their ways to even consider them. A major problem in the introduction of new foods is not the food itself, but with the people altering their way of thinking and accepting the food, which although different and alien to them, might often be equally nutritious and capable of being produced less expensively and more abundantly.

The Indians, though they didn't grow them all as crops, utilized many of our native plants. In 'The First Canadians' (Symington, 1978), it is stated that:

"He 'tJi 11 make hemlock tea to prevent scurvy - hemlock having a high Vitamin C content. This is not necessary now because his diet includes rose hips, fish entrails, cattail hearts and root nodes, and various other herbs including the mint with which he heightens the flavour of meat .broth and stew, and which he

dries for aromatic tea. 11

There is a great abundance of wild edible foods all around us (see Table 2), rarely utilized except by the occasional nature and health enthusiast.

- 5 - TABLE 2 SOME WILD EDIBLE FOODS PLANTS PART UTILIZED Dandelions Leaves Salad, Vegetable - rich in Vitamin A Flowers Wine Roots Caffeine-free Coffee Wild Rose Hips Raw - tea, soup, jelly Sweet Violets Flowers Wine, Desserts, Broths, Salad - very rich in Vitamins A & C Leaves Vegetable, Salad Dock Seeds Flour Leaves Salad, Vegetable - more Vitamin c than oranges, more Vitamin A than Carrots Clover Leaves Vegetable Flowers ~~ine Wild Rice Seed A Gourmet Delicacy Cattails Roots As Potatoes, Flour Flowers Flour Stalks Boiled when Young Sassafras Leaves Tea, Soups, Salads, Jelly - much Vitamin C Water Lily Seeds Popcorn, Breakfast Cereal Roots as Potatoes, Flour Leaves Boiled Several Times Spruce Trees Inside Bark Jack in the Pulpit Roots Skunk Cabbage Roots Milkweed Stalks, Blossoms Sunflower Seeds

- 6 - TABLE 2 (continued)

Goldenrod Leaves Tea Wampee, Bur Reed Seed 11 Swamp Corn 11 Prickly Lettuce Watercress Calamus (Sweet Flag) Leaves, Stalks Roots Tea, Candy Sheep Sorrel Giant Puffball Silverweed Slippery Elm Inner Elm Bark Boiled (River Elm)

- 6 A - YELLOW WATER LILY (NUPHAR ADVENA)

Nuphar is a genus of about seven species of aquatics, native in the temperate regions of the Northern Hemisphere. They grow in ponds, slow-running streams, and swamps. Nuphar Advena, the Yellow Water Lily~ also known as Common Spatterdock or Cow-Lily, is found throughout the eastern half of the U.S.A. and Canada. It flowers from June to August, with large globular flowers averaging two to four inches across. Centered in each flower are numerous reddish stamens, bright in season with yellow pollen. Later in the summer the flowers develop large seeds resembling kernels of corn (Outdoor Canada 5(7)). The leaves are large, erect, and heart shaped. The rootstalk, scaly and slightly yellowish, resembling banana stalks, may be as much as four or five inches thick and 10 feet long.

The roots are eaten by black bear, beaver, muskrat, and moose; and contain large amounts of starch. Indians boiled and roasted the roots as a vegetable. John Josselyn, one of the first chroniclers of New England natural history, recounts that:

11 The Indians Eat the Roots, which are a long a

boiling, they taste like the Liver of a Sheep 11 while Or. Edward Palmer stated that, although the squ~wsoften dove for these rootstocks, they found it simpler to steal them from the muskrat houses in \vhich they were stored (Fernald & Kinsey, 1958).

The seeds are eaten by mallards, cranes, and rails. The Indians used the seeds roasted or ground them into meal for bread. Popcorn can be made from the seeds, or they can be eaten as a breakfast cereal.

- 7 - The main plant (stalk ~nd leaves) was eaten by the Indians, and also by beavers, muskrats, and porcupines. Recent research has been done on the chemical composition of Water Lily leaves and petioles, but none appears to have been done on the roots.

Reimer and Toth (1970) did mineral analyses on the leaves and petioles of spatterdock and four other species of nymphaeaceae. Boyd (1968) determined the chemical composition of dried Water Lily plants to evaluate their potential as roughages. No data is available on the roots or flowers of the Yellow Water Lily.

There have been some studies on cattail roots, the closest plant to the Water Lily and for which there is data on roots. Cattail rootstocks contain in winter 12.5 parts starch to 73 parts water; in spring, only 10.5 parts starch to the same quantity of water. Analyses of defibered cattail rootstock flour have shown:

Moisture 7.35 - 8.78% Ash 2.48 - 2.84% Fat 0.61 - 4.91 % Protein 7.22 - 7.75% Carbohydrates 79.09 - 81.41 %

(Morton, 1975)

- 8 - AQUATIC PLANTS - PESTS

Aquatic plants interfere with transportation, water sports, and fisheries. They decrease the rate of flow of water, increase evaporation losses, and many aquatic weeds harbour specific insect vectors of human and animal dieseases (Gupta~ 1973; Heffron et al~

1977). In Florida in 1970, more than one ~illion doll ars were spent on partially effective efforts to keep its 4000 square miles of infested waters free of aquatic weeds. In the developing countries of the world, the loss of potential gross national product is proportionately larger than in the United States. It has been estimated that direct economic losses to these areas exceed lOO million dollars annually (Bates & Hentges, 1976).

Methods of aquatic weed control include biological, chemical, and mechanical.

Nutrients which feed aquatic plants are leached from the soil or derived from sewage run-off, and are ultimately destined to promote undesirable growth of some kind, or to reach the oceans without being utilized but with their damaging potential intact. The weed plants themselves represent a considerable photosynthetic effort in which these dilute nutrients, unavailable to other flora, together with atmospheric carbon dioxide and solar energy, are efficiently concentrated (Bates & Hentges, 1976).

Harvesting and processing aquatic plants as a source of methane for fuel and protein has been studied and reviewed. A few

- 9 - investigators have studied nutrient availability in aquatic plants in combination with dry pelleted cattle rations (Heffron, 1977).

Although aquatic vegetation, particularly certain types of seaweed, have valuable primary uses for food in the Orient and secondary uses as industrial stabilizing agents in processed foods, aquatic plants are not widely consumed as human food. The negative aspects include high moisture content, fibrous bulk, unappetizing appearance, off-flavors, poor digestibility, and occasionally toxic components. With few exceptions, human feeding goals have centered upon the recovery and purification of leaf protein (Bates & Hentges, 197 6).

The major problem which would be involved in harvesting

~Jater Lily roots for food would be the handling of a bulky crop at a high moisture content. However, in industrialized countries, automation is a possibility. In poorer countries, labour intensive procedures would be most efficient.

- 10 - FERMENTATION

Fermentation and drying are the oldest methods of food preparation known to mankind. Fermentation is a method of preparing food in order to develop certain desirable characteristics. The changes in flavour, aroma and. texture are as unique for each food as the masterpieces of the world renowned cooks (Carr et al, 1979).

From the biochemical standpoint, fermentation if the name given to the general class of chemical changes or decompo~itions produced in organic substrates through the activities of living microorganisms.

The microorganisms of fermentation include yeasts, molds, and bacteria. These microorganisms are unable to manufacture their own food by the ordinary process of photosynthesis since they lack chlorophyll. The microorganisms of fermentation differ widely in respect to morphology, size, reaction to free oxygen, manners of reproduction, growth requirements, ability to assimilate or ferment raw (natural) substances, and in other ways. But they are similar in that they are 'colorless' and grow most actively in darkness or diffused light, and all produce enzymes by which they catalyze the reactions ascribed to them (Prescott & Dunn, 1949).

There are numerous fermented foods made throughout the world, mainly in the Orient. These include Miso from Japan (fermented soybeans and rice), Shoyu from Japan (fermented soybeans and roasted wheat or wheat flour), and Tempeh from Indonesia (fermented soybeans). rp Th~most l e der son, 1971)·=: commonly known in the western world is sauerkraut or

- 11 - I fermented cabbage. Fermentation is also involved in the pickling process of cucumbers, green tomatoes, beans, beets, etc.

A variety of wild plants have been reported to be used in the past for pickling. Included are the roots of wild onion, cattail, false spikenard, Solomon's seal, Indian cucumber, live-forever, bugleweed, Jerusalem artichoke, succulent leafy or young plants for fleshy branches of sapphire, pokeweed, sea-pursline, sea-milkwort, and flowerbuds of marsh marigold, barberries, redbud, elder buds, and ash buds. These have usually been placed in salt solutions and later packed with weak vinegar. There is little scientific evidence to corroborate such reports (Pederson, 1971).

Lactic acid bateria are characterized by their ability to convert glucose, and other sugars into lactic acid when grown on suitable media. In nature, lactic acid bacteria chiefly occur on plants and vegetable matter. Some also occur on the mucous membranes of man and animals. Most of the spontaneous souring of food, especially milk, is due to lactic acid bacteria.

The lactic acid bacteria are extensively employed in dairying and their application in this industry is the most important, but in other fields of industry, the ability of lactic acid bacteria to produce considerable quantities of an organic acid is utilized. The acid is added to foodstuffs for the purpose of imparting a special flavour or for improving its keeping qualities (Jorgensen, 1948).

Bacillus Cucumeris and Bacillus Brassicae are involved in nearly all vegetable fermentations. Leuconostoc Mesenteroides

- 12 - initiates sauerkraut, and is involved in the fermentations of b.eets, cucumbers, turnips, sliced green tomatoes, cauliflower, etc. Lactobacillus Plantarum is a high acid-producing bacteria, and plays a major role in the fermentation, particularly in brines. Lactobacillus Brevis is important to imparting character to fermented vegetables (Pederson, 1971).

A large number of carbohydrates may be utilized for lactic acid production. Corn and potato starches may be hydrolyzed by enzymes to maltose and glucose. The lactic acid fermentation is generally carried out at a comparatively high temperature (30 - 50 C depending on the bacteria utilized). The sugar in the mashes is normally adjusted to a concentration of 5 to 20% depending on the raw material and conditions of the process. The fermentation proceeds best in slightly acidic conditions (Prescott & Dunn, 1949).

SAUERKRAUT

Cabbage contains:

86 - 94.3% Water 2.9 - 6.4% Sugar 0.2 - 2.4% Protein 0.1 - 1.6% Fiber and 0.4 - 2.4% Ash.

The three lactic-acid-producing bacteria types that develop in the normal controlled fermentati on are Leuconostoc Mesenteroides (@ salt = 2. 5%, temp. = 70 F), then Lactobacillus Cucumeris and L. Plantarum, and finally L. Pentoaceticus (L. Brevis). A salt concentration of 2.5% - 13 - I is usually used, and the optimum temperature is 65-70 F. It is necessary to exclude oxygen (and therefore yeasts and spoilage types of bacteria) ~y covering the sauerkraut (Prescott & Dunn, 1949).

- 14 - EXPERIMENTAL WORK

- 15 - COLLECTION AND TREATMENT

Two large bagfuls of Water Lily roots were collected from Three Lakes in the Eastern Townships of Quebec. The roots grew near the shore in water two and a half feet deep. The mud was extremely soft, and collection was most easily done by hand digging. The roots of Water Lilies are long, with several plants growing off each root, resulting in considerable root material to harvest in an area.

The roots were transported in lake water, minimizing dehydration and leaching. They were then washed, peeled, cubed (1 - 1~"), blanched in boiling water for approximately three minutes, packed, and then frozen in plastic bags with a 220 gram content each. The total harvest was approximately five kilograms.

Two bags of roots were freeze dried (!" cubes) for 24 hours at -50 C and 10 fL pressure, for use in future analyses.

- 16 - FERMENTATION

All samples were fermented in 16 ounce mason jars in darkness. An attempt to keep the fermentations anaerobic by maintaining the liquid level above the solids, and by covering the surface with plastic wrap was done in all cases. Two trial and two batch fermentations ·were done.

pH readings were taken approximately every 24 hours, and a sample was considered to have finished fermenting when the change in pH was less than 0.1 units. Acidity also was measured by means of titration with sodium hydroxide (see calculations, following page).

The first trial fermentation was a jar of cubed roots in a 3% brine at 20 C, innoculated with bacteria from a sauerkraut fermentation. This run was unsuccessful because the initial pH \vas 4.3, dropped to 3.9 in 24 hours, then stagnated. Lactic acid bacteria stops growi·ng at a pH of around 4; therefore, a good starting pH is about 6.

The second trial fermentation consisted of two jars. Shredded roots, as opposed to cubed were used in order to expose more root surface to the bacteria, and to make nutrients more available. The pH in both jars was brought to 6 by the addition of sodium hydroxide. The samples were innoculated with lactic acid bacteria growing on malt sprouts in a nutrient solution. The first jar had a 3% salt and a 3% sugar content, the second jar had a 3% salt content with no sugar (see recipe).

- 17 - CALCULATIONS FOR ACIDITY OF SAMPLE

Normality of NaOH = 0.01619 (as titrated against potassium acid thalate, KHC8H4o4) Indicator used= phenolphthalein ·

Temperature = 20 c

Acidity (mg CaC03/liter = (ml NaOH) x (NNaOH) x (50000) ml sample

Batch # 2 Fermentation

Initially: 2.5 ml sample takes 1.0 ml NaOH

acidity= (1.0) (0.01619) x x (50000) = 324 mg CaC0 3/liter 2.5

Finally: 2.0 ml sample takes 1.25 ml NaOH

acidity= (1.25) x x (0.01619) (50000) = 506 mg CaC03/liter 2.0

- 18 - R E C I P E

lOO grams Water Lily Roots

50 grams water

6 grams salt (for a 3 % salt concentration) (4 grams for a 2 % concentration)

* 6 grams sugar (for a 3 % sugar concentration)

NaOH to bring pH to 6

Water to bring weight to 200 grams

* Determined not to be necessary.

- 19 - The samples took six days at 20 C to lower their pH's to 4.6 ' and 4.75 respectively, where they stagnated. There was no difference in the rate of fermentation of the sugar and sugarless sample (see Table 3 and Figures l & 2). A microscope slide taken from each sample showed that the sugarless sample contained no lesser a bacteria count than the sugar sample (in fact it was a larger count, though this cannot be considere~ significant since only one sample was done). Therefore, it was concluded that the sugar was unnecessary in the fermentation.

In the first batch fermentation, 12 jars of shredded Water Lily roots were innoculated with malt sprout bacteria. Six samples had 2% salt content, with 3 of them fermenting at 20 C, and 3 at 25 C. The remaining six had a 3% salt content, with 3 at 20 C, and 3 at 25 C. · The samples all took 5 days to get to their final pH. All six of the 3% salt samples molded, while only one (at 25 C) of the 2% salt samples molded. A possible explanation would be that the 3% salt inhibited the yeasts and bacteria, thus decreasing the competition for mold cells, and making it easier for the mold to grow. From these results, it was decided to use a 2% salt concentration for the final batch fermentation. The unmolded 2% salt concentration fermented roots were tested on mice (see section on the mice trial).

On the final (#2) batch fermentation, ten jars of shreddeo roots at a salt concentration of 2% and a pH of 6 were innoculated with malt- sprout lactic acid bacteria. The fermentation took 3 days at 20 C. Slides were taken at innoculation, and at the end of the fermentation to inspect the bacteria. Yeast and nutrient augers were taken from one - 20 - of the samples to determine the microorganisms ptesent. No molds were observed, but lactic acid bacteria and some yeasts were present.

The acidities (as calculated by the NaOH titration) were 324 mg CaC0 /liter initially, 3 and 506 mg CaC03/liter finally (see Calculations, Page 18).

The samples from this fermentation were used for the taste panel, and the content analyses.

- 21 - TABLE 3

TRIAL SAMPLES NO. 2 ·

SAMPLE 1 (sugar) SAMPLE 2 (no sugar)

·Eli Acidity (mg CaC03/l) £!!_ Acidity

0 hrs 6.0 6.05

23 hrs 5.5 526 5.65

53 hrs 5.25 5.25

70 hrs 5.15 742 5.15 567

95 hrs 4.95 5.0

118 hrs 4.80 854 4.85 876

142 hrs 4.6 4.75 *finished

167 hrs 4.6 1619 4.75 1012

- 22 - FIGURE 1

o sugarless sample • 3 °/o sugar sample 6.1

:r: a. 5 .I

0 24 48 72 96 120 144 168 192 TIME ( hrs )

SA~1PLE 2

pH vs time for the sugarless and 3% sugar samples.

- 23 - FIGURE 2

1900 o sugarless sample a 3 °/o sugar sample ,... 1700 .._ .....(\) • = 1500 ...... ,. 0 slope : 8 1300 7.1 intercept = 265 CJ) correlation ' 0.923 -E 1100

>-.,.._ 900 slope • 3.5 -Cl intercept • 419 u 700 cor relation : 0.954 <(

0 500

300

0 24 48 72 96 120 144 168 192 TIME ( hrs )

SAMPLE 2

Acidity vs time for the sugarless and

3% sugar samples.

- 24 - MOISTURE CONTENT

Moisture content was determined by weighing fre~h samples, then drying them in an oven at lOO C for 24 hours or until all the water had evaporated (i.e., therewasno further change in weight). The percent water content ·was obtained by dividing the change in weight by .the total initial weight (wet basis).

For the fresh straight-from-the-lake Water Lily roots, 10 samples were dried. The average moisture content was found to be 86.5%, with a standard deviation of 1.6%. (See Table 4)

The fermented Water Lily roots were saturated, as they were covered by liquid in the mason jars. Ten samples were dried, and the average moisture content found to be 92.4%, with a standard deviation of 0.1 %. (See Table 4)

- 25 - TABLE 4

MOISTURE .CONTENT (Wet Basis)

FRESH SAMPLES FERMENTED SAMPLES SHREDDED SAMP~ES No. Moisture No. Moisture No. Moisture Content (%) Content (%) Content (%)

1 88.48 1 92.47 T2 88.8 2 88.68 2 92.42 Bl 8.9.5 3 87.36 3 92.29

4 86.83 4 92 . 26 5 86.32 5 92.27 6 86.00 6 92.25

7 85.91 7 92.63 8 86.52 8 92.35 9 86.15 9 92.38 10 83.03 10 92.21

- 26 - ASH CONTENT

Total ash (mineral) content was determined by weighing the samples, then burning them in a muffle oven at 500 C for an hour or until there was no further change in weight. Upon removing the samples from the oven, they ·were immediately placed into a dessicator to cool until weighing, ensuring that they absorbed no moisture before weighing.

Five samples of freeze-dried roots were burned, and the average ash content was found to be 3.83% of the dry matter, or 0.52% of the total matter. (See Table 5)

Five samples of fermented roots were burned, and the average ash content was found to be 29.50% of the dry matter, or 2.26% of the total matter. (See Tab 1e 5)

- 27 - TABLE 5

ASH CONTENT

Unfermented Samples Fermented Samples No. Ash( % D.M.) Ash( % total Number) No. Ash( % D.M.) Ash( % T.M.)

1 4.099 0.533 1 30.218 2.312

2 3.518 0.475 2 29.104 2.226

3 3.552 0.480 3 29.569 2.262

4 4.380 0.591 4 29.014 2.219

5 3.592 0.485 5 29.589 2.263

Ash (% T.M.) = Ash (% D.M.) x (100%- % Moisture Content) = Ash (% D.M.) x % D.M.

- 28 - PROTEIN CONTENT

Total protein content-was determined by the Kjeldahl method which requires a sample size containing at least 3 mg N for accurate 2 results.

The sample must ·be accurately weighed, then digested (boiled) with sulfuric acid and KEL-PAK #5 which contains potassium sulfate and mercuric oxide. The potassium sulfate increases the digestion temperature, and the mercuric acid encourages the sulfuric acid to oxidize. As the sulfuric acid oxidizes, it combines with the organic nitrogen to form ammonium compounds in the mercuric acid.

When the digestion is finished, a NaOH-thiosulfate solution is added, and a steam distillation carried out. The sodium thiosulfate frees the ammonia, which is condensed into a boric acid solution and subsequently titrated with sulfuric acid. The sodium hydroxide provides the required alkaline medium for distillation. The nitrogen content is then calculated, and the protein determined by multiplying nitrogen content by 6.25.

Five freeze-dried root samples were done and an average protein content of 5.18% of the dry matter (with a standard deviation o·f 0.44%) was found, or 0.70% of the total matter. (See Table 6)

Five oven-dried fermented samples were done and an average protein content of 4.60% of the dry matter (with a standard deviation of 0.05%) was found, or 0.36% of the total matter. (See Table 6)

- 29 - TABLE 6 PROTEIN CONTENT

Unfermented Samples Fermented Samples No. Protein (% Dry Matter) No. Protein (% Dry Matter)

1 4.90 1 4.67 2 4.83 2 4.73 3 5. 31 3 4.64 4 4.88 4 4.77 5 5.98 5 4.65

CALCULATIONS

%N 2 = (ml HCl) x (14.008) x (100) x (NNaOH) (1000) x (g sample}

NNaOH = 0.01619

% Protein = 6~25 x % N2

Protein (% Total Matter) = Protein (% Dry ~1atter) x (100% - % Moisture Content) = Protein (% Dry Matter) x % Dry Matter

- 30 - FAT CONT~NT

Fat content· was determined by the ether extraction method. The sa~ples were weighed, then hot ether (which extracts the fat) was percolated through the dry ground samples for 24 hours. The samples were then dried and reweighed, and the difference in weight is recorded as the fat content.

Two samples of five grams each of the freeze-dried roots were analyzed and the fat content found to be .62% (D.M. basis).

Two samples of five grams each of oven-dried fermented· roots were analyzed and the fat content found to be .30% (D.M. basis).

The results of the complete content analysis can be seen. in Table 7.

- 31 - TABLE 7

TOTAL CONTENT ANALYSlS

Fresh Water Lily Roots Fermented Water Lily Roots

% of D.M. % of W.M. % of D. M. % of ~1. M.

Water 86.5 92.4

Ash 3.83 0.52 29.50 2.26

Protein 5.18 0.70 4.60 0.36

Fat 0.62 0.08 0.30 0.02

Carbohydrates 90.4 12.2 65.6 5.0

- 32 - . MICE . TRIAL

Eleven mature .male albino rhine mice with identical traits and characteristics, upon which the fermented Water Lily roots were to be tested~ were purchased.

Six of the mice were fed "x" amount of mouse feed combined with "x" amount of batch #1 fermented Water Lily roots, and water. The remaining five mice (the controls) were fed "x" amount of mouse feed with water to bring it to the same consistency as the test feed. In this manner, the test mice were fed the same amount of mouse feed as the control mice, plus the fermented Water Lily roots. The mice followed their respective diets for 8 days, having unlimited food made available to them (both groups ate equal amounts of mice feed, not including fermented roots). During this period, the six test mice ate 300 g total of ferm~nted Water Lily Roots. As this is 50 g each, and the mice weighed only 28.4 g (average) initially; it was concluded that the fermented Water Lily r.oots contained no toxic components in d~ngerous amounts.

An interesting phenomena was that the test mice gained an average of 0.3 g each, while the control mice gained an average of 3.1 g each (a gain 10.3 times greater). This could be due to the presence of some component in the fermented Water Lily root which inhibits the digestion and breakdown of food. (See Table 8)

.., 33 - TABLE 8

MICE TRIAL

Initial Weight (g) Final ~Iei ght (g) Weight Gain (g)

"TEST MICE

# 1 27.2 28.1 0.9

# 2 28.7 29.7 1 0 0

# 3 27.1 27.8 0.7

# 4 27.9 26.8 - 1 . 1

# 5 28.4 30.1 1 . 7

# 6 31 . 1 29.8 - 1. 3 AVERAGE 28.4 28.7 0.32

CONTROL f~I CE

# 7 30.3 34.2 3.9

# 8 29.3 33.0 3.7

# 9 31 . 1 33.9 2.8

# 10 28.2 31.0 2.8

# 11 28.6 31 . 1 2.5 AVERAGE 29.5 32.6 3.14

- 34 - TASTE PANEL

A taste pane.l was conducted on the fermented Water Lily roots of batch #2. The fermented roots were heated and mixed half and half with mashed potatoes, then served hot with a touch of salt and pepper. Sauerkraut was prepared in the same manner as a control, and the panelists had to judge the two samples without knowing which was the fermented roots. The tests were conducted under red lights to hide the pinkish-gray colour of the roots-potatoes mixture. In this way, it was hoped that judgements would not be influenced by appearance. Each panelist was given enough sample for more than one taste if so desired

(th~ugh not many so desired), and each panelist was seated at his/her own cubicle where he/she would not be influenced by the reactions of others.

The questionnaire consisted of three questions, and room for comments. "Do you consider this an acceptable food?" is a standard question to obtain the overall opinion of the sample. "If you could buy this product at 50 t /lb. or peas at $5 I lb., which would you eat? (assuming comparable nutritive value)" was included to determine if people would eat this sample, given a definite economic advantage, or if this sample was so bad that it is not worthwhile to examine it as a food source, no matter what the economic advantage. "Which food do you prefer?" questionned whether the fermented vJater Lily roots were preferable over a similar (sauerkraut) accepted food.

A total of 24 panelists participated. The results can be seen in Paae_. 38. 33% of the panelists found the fermented Water Lily - 35 - roots an acceptable food and would eat them ratner than peas at the stated prices; as opposed to 77% for the sauerkraut. There were several comments that the Water Lily sample was too bitter, or that it had a bitter aftertaste with the first taste being palatable but the second taste bitter. It was suggested that it might be better tasting if prepared in a different manner with an addition to decrease the bitterness; i.e., spices. The fermented Water Lily roots were also

11 described as "the pits" and "yuk! •

- 36 - QUESTIONNAIRE FOR TASTE PANEL EVALUATION

SAMPLE 1 SAt~PLE 2

Do you find this an acceptable food? Yes No Yes No

If you could buy this product at 50 i/lb. or peas at $5/lb., which would you eat? (assuming comparable This Peas This Peas nutritive value) Sample Sample

Which food do you prefer? This This One One

Additional Comments: ......

- 37 - TASTE PANEL RESULTS lo Do you find this an acceptable food?

Fermented Water Lily Food Yes: 8 No: 16

Sauerkraut Yes: 19 No: 5

2. If you could buy this product at 50 t I lb. or peas at $5 I 1b. , which would you eat? (assuming comparable nutritive value)

Fermented Water Lily Root: 8 Peas: 16

Sauerkraut: 18 Peas: 6

3. Which food do you prefer?

Fermented Water Lily Root: 2 Sauerkraut: 22

- 38 - DISCUSSION AND CONCLUSIONS

Water Lily roots were not found to be very nutritive, either fermented, nor unfermented. The protein content of the unfermented Water Lily roots is low, comparable to that of cucumbers and muskmelons (see Table· 1), and lower than that of cattails. The protein content of the fermented Water Lily roots is lower than any of our conventional vegetables. In comparing the ash content, the unfermented sample is comparable to other vegetables, while the fermented sample is at the

~igher end of the ash content range.

The results of the taste panel show poor consumer acceptance of the product, and this, coupled with a poor nutritive value, makes it doubtful that Water Lilies are suitable for cultivation for this purpose.·

However, in situations where the Water Lilies must be cut as an aquatic weed or in places where the acreage could have absolutely no other use and there is a shortage of feed, it might be worthwhile to harvest the roots as a silage feed for animals. There would be extensive problems to be considered however. The roots would have to be partially dehydrated before ensiling. It would have to be determined if a machine which harvested the roots would be economically feasible and possible. Study would also have to be given to the reason why the test mice which were fed fermented Water Lily roots did not gain weight as rapidly as those which were control mice as this might cause problems when trying to fatten up pigs and beef cattle.

Fermented Water Lily roots might also prove to be an acceptable diet food in that one may eat all the food which one desires so long - 39 - as it is accompanied by an equivalent amount of fermented' Water Lily roots, and thus one will gain no weight.

- 40 - BIBLIOGRAPHY

- 41 - BIBLIOGRAPHY

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Boyd, C.E. 1968 Fresh Water Plants : A Potential Source of Protein Ec. Bot. 22 pp 359-368

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Couvillon, Carolyn · 1978 The Mealworm - A Potential Source of Protein Agricultural Engineering Senior Project

Delman, D. 1978 Nature's Pantry International Wildlife 8(5) pp 28G 8(6) pp 320

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Gupta, Or. 0. P. Aquatic Weed Control World Crops J-A 1973 pp 182-189

Hedrick, U.P. 1933 A History of Agriculture in the State of New York New York; J.B. Lyon Company

Heffron, C. L. et a 1 1977 Chemical Composition and Acceptability of Aquatic Plants in Diets of Sheep and Pregnant Goats J. Anim. Se. 45 pp 1166-1172

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- 43 - House, H.D. 1935 vJi l d Flowers New York; Macmillan

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Parrish, G. K.; Kroger, M.; Heaver, J.C. 1974 The Prospects of Leaf Protein As A Human Food CRC Critical Reviews in Food Technology 5 pp 2 - 44 - Pederson, C. S. 1971 Microbiology of Food Fermentations Westport, Conn.; The AVI Publishing Company, Inc.

Pirie, N. W. 1975 Food Protein Sources Cambridge; Cambridge University Press

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- 45 - Water Lilies - Produce From the Pond 1977 Outdoor Canada 5 (7) pp 8-9

- 46 -