Biology 3460 Weeks 2, 4, 6, & 8

Biology 3460 - Plant Physiology - Lab Exercise 3

Plant Mineral Nutrition
Objectives:
This lab exercise is intended to:
(1) introduce students to the mineral nutrient requirements of plants,
(2) illustrate sand culture as a technique to grow plants under controlled nutrient
conditions,
(3) provide experience making plant nutrient solutions and calculating nutrient
concentrations in different solutions,
(4) test of the effect of different sources of nitrogen on growth of grasses.

Introduction
Mineral nutrition is the study of how plants obtain and use basic mineral nutrients
normally taken up by roots from the soil solution. Essential mineral nutrients required by
plants are shown in Table 5.1 in your textbook (attached). Access to these nutrients in
sufficient supply, along with adequate water and sunlight allows plants to synthesize all
the other components they require for growth. An empirical formulation of a complete
nutrient solution for plants was developed by Dennis Hoagland, a researcher in
California, USA. There have been several modifications of Hoaglands solution over the
years with a typical example of a modified Hoaglands solution illustrated in Table 5.3
of your textbook (attached). Hoaglands solutions typically provide nutrient elements in
ratios related to the nutrient requirements by plants, but the nutrient concentrations in full
strength modified Hoaglands solution can be several times higher than a plant would
experience in the natural world. When growing non-agricultural plants it is often
necessary to dilute the modified Hoaglands solution in order to prevent injury due to
toxicity or salinity stress, particularly with young seedlings.
Typical modified Hoaglands solutions provide nitrogen as both ammonium (NH4+) and
nitrate (NO3-) ions. Most plants grow best when nitrogen is supplied from both
ammonium and nitrate. If nitrogen is supplied as only one type of ion (either nitrate or
ammonium) large changes in the pH of the nutrient solution can occur and this can then
disrupt the uptake of other required nutrients such as calcium, magnesium or iron. Plants
generally grow better when they have access to both ammonium and nitrate because this
promotes cation-anion nutrient balances within the plant.
Instructions for making the macro-nutrient stock solutions for modified Hoaglands
solution are shown below. Three different formulations are illustrated, one with both
ammonium and nitrate supplied (typical Hoaglands solution) and then two solutions in
which nitrogen only in the form of either nitrate or ammonium is supplied. In this lab
experiment, plants will be grown in one of three treatments (both ammonium and nitrate,
nitrate only, and ammonium only) to test the effect of nitrogen source on plant growth
(biomass (dry weight) production). Note that it is not possible to keep the concentrations
of all macro-nutrients at identical concentration in all three versions of the modified
Hoaglands solution (Calcium concentration is modified in the nitrate-only solution and
Sulphur concentration is modified in the ammonium-only solution).

Biology 3460 Weeks 2, 4, 6, & 8

In addition to the macro-nutrients shown in the nutrient solution recipes illustrated below,
the modified Hoaglands solution also provides micro-nutrients from a prepared stock
solution and iron from a chelated-iron solution, which keeps the iron in a soluble form
and prevents it from precipitating out of solution and making it unavailable to the plants.
Part A. The Nutrient Solutions
Modified Hoaglands Solution (both NO3 and NH4 provided)
Macro-nutrient Stock Solutions:
1M Calcium nitrate [Ca(NO3)24H2O]
1M Potassium nitrate [KNO3]
1M Magnesium sulphate [MgSO47H2O]
1M Ammonium phosphate [NH4H2PO4]
To make 4 litres, add:
4 ml calcium nitrate
6 ml potassium nitrate
2 ml magnesium sulphate
6 ml ammonium phosphate
2 ml iron + 2 ml micronutrients
Top up to 4 L with distilled water
Modified Nutrient Solution (only NO3 provided)
Macro-nutrient Stock Solutions:
1M Calcium nitrate [Ca(NO3)24H2O]
1M Magnesium sulphate [MgSO47H2O]
1M Potassium phosphate (monobasic) [KH2PO4]
To make 4 litres, add:
10 ml calcium nitrate
2 ml magnesium sulphate
6 ml potassium phosphate
2 ml iron + 2 ml micronutrients
Top up to 4 L with distilled water

Biology 3460 Weeks 2, 4, 6, & 8

Modified Nutrient Solution (only NH4 provided)

Macro-nutrient Stock Solutions:
1M Calcium chloride [CaCl22H2O]
1M Potassium phosphate (monobasic) [KH2PO4]
1M Magnesium sulphate [MgSO47H2O]
1M Ammonium sulphate [(NH4)2SO4]
To make 4 litres, add:
4 ml calcium chloride
6 ml potassium phosphate
2 ml magnesium sulphate
10 ml ammonium sulphate
2 ml iron + 2 ml micronutrients
Top up to 4 L with distilled water
Protocol:
Work as individuals to complete this part of the lab.
1. Calculate the concentration (mmol/L) of each of the macro-nutrients for the three
modified Hoaglands solutions and fill in the table below. A sample calculation is
provided.
Sample Calculation:
For the Hoaglands solution with both ammonium and nitrate present, the concentration of
potassium (K+) is determined as follows:
6 ml of 1 M potassium nitrate was added to a bottle and the volume is made up to 4 L.
6 ml x 1 mmol/ml (1 M) potassium nitrate = 6 mmol of potassium nitrate
6 mmol potassium nitrate in 4 L of solution is: 6 mmol/4 L = 1.5 mmol/L
Table 1. Concentration of individual macronutrients available in three modified Hoaglands
solutions.
Macro-Nutrient Both NO3- & NH4+
NO3- only
NH4+ only
(mmol/L)
(mmol/L)
(mmol/L)
N
P
K
Ca
Mg
S

Biology 3460 Weeks 2, 4, 6, & 8

Part B. The Experimental Design

Plants will be grown in sand culture in each of the three nutrient treatments, under the same
light conditions. We will measure total dry biomass of the plants at the end of a four-week
growing period.
Protocol:
Work as a group of three students to complete this part of the lab.
1. Send one of your group members to help with mixing the first batches of the three
nutrient solutions, according to the recipes outlined in Part A.
2. Prepare one plant pot for each of the three treatments (both NO3 and NH4, only
NO3, and only NH4). Be sure to label the pots with the treatment type and a group
identifier. Add a layer of polyfilter wool to the bottom of the pot to prevent sand
from escaping through the holes. Fill the pot with sand up to about 2 cm below the
lip of the pot.
3. Several grass seedlings (your instructor will inform of the species used) have been
germinated in Petri dishes in Promix soil mixture. Carefully choose four seedlings
for each treatment (will need 12 seedlings in total), be sure to choose healthylooking seedlings of about the same stage of germination. Remove the seedlings
very gently from the Petri dish, and rinse them under a gentle stream of water to
remove as much soil from the roots as possible.
4. Moisten the sand in the prepared plant pots with the appropriate nutrient solution,
then make wells in which to plant the four seedlings for each treatment. Poke a
well with your finger deep enough to support the roots of the seedling, but not so
deep that the shoot will be under sand once planted. Place the seedling into the well
then gently push sand around it to fill in the well and support the seedling. Be sure
to space the four seedlings evenly throughout the pot.
5. Once you have completed a pot for each treatment, place them in the appropriately
labeled plastic tub. Your lab instructor will fill the tubs with the nutrient solutions
prepared in Step 1. Nutrient solutions will be changed twice a week. You may be
asked in future labs to mix fresh nutrient solutions and refill the tubs.
6. At the end of the four-week growing period, cut the plants off the roots right at the
surface of the sand, place them in an envelope for each treatment (label your group
names and the treatment type on the envelope). The above-ground plant material
will be dried in a drying oven for two weeks. In the next lab period we will collect
class data for the above-ground biomass for each treatment.

Biology 3460 Weeks 2, 4, 6, & 8

Treatment
Bench 1
Both NO3- &
NH4+
NO3- only

Bench 2

Above-ground Biomass (g)

Bench 3
Bench 4
Bench 5

Mean
SD

NH4+ only

Analysis:
1. Use class results to prepare a figure that shows the effect of mineral nutrition treatment
on the above-ground biomass of the plants. Mean biomass values with standard
deviations should be plotted.
2. Using a Kruskal Wallis Test, determine if there is a statistically significant difference
between the treatments.
3. What effect did the different nutrient regimes have on plant growth? Why? Explain.

Biology 3460 Weeks 2, 4, 6, & 8

Biology 3460 Weeks 2, 4, 6, & 8