Protocol

Author

Rana Munns

Overview

This protocol describes how to make up standard nutrient solution using the original recipe of Hoagland and Arnon No2, with modifications for using it in solution culture rather than sand culture.

Background

Growing plants under defined nutrient availability has been useful for nutrient studies and for screening for a variety or nutrient deficiencies and toxicities. The original solutions of Hoagland and Arnon were for sand culture, which need to be modified for solution culture for P.

Materials/Equipment

• 2 L plastic beaker
• 2 L volumetric flask
• Magnetic stirrer
• Electronic balance which measures to 4 decimal places.
• Funnel
• 1 L Schott bottle

Units, terms, definitions

All concentrations are given in mM (mmol L^-1or mol m^-3)

Procedure

Make up stock solutions A (containing K, Ca and NO₃), B (containing Mg, NH₄and PO₄), C (all micronutrients) and D (FeCl₃). These cannot be mixed at the high concentrations of the stock solutions.

Stock	Compound	Final concentration	MW	Amount of compound per L	Stock concentration (mM)	Volume per L to add (mL)
A	KNO3	6.5 mM	101.11	82.15	812	8
	Ca(NO3)2.4H2O	4.0 mM	236.16	118.08	500
B	NH4H2PO4	2 mM	115.03	28.8	250	8
		0.1 mM	115.03	1.44	12.5
	MgSO4.7H2O	2.0 mM	246.47	61.62	250
C	H3BO3	4.6 M	61.83	0.284	4.6	1
	MnCl2.4H2O	0.5 M	197.9	0.099	0.5
	ZnSO4.7H2O	0.2 M	287.54	0.055	0.2
	(NH4)6Mo7O24.4H2O	0.1 M	1235.95	0.124	0.1
	CuSO4.5H2O	0.2 M	249.7	0.050	0.2
D	FeCl3	45 M	162.2	24 ml of 60%	45	1

When using solution culture rather than sand culture, for which the original Hoagland and Arnon solution was devised, it is advisable to reduce the concentration of PO₄to low levels to avoid P toxicity due to excessive uptake of certain species such as lupin and wheat. This means that P uptake should be monitored and P added regularly.

To make stock solution A, weigh both ingredients into a 2 L plastic beaker and top up with distilled H₂O to around 1.8 L. Place stirrer flea in container and place on stirrer for a few minutes until all the ingredients have dissolved. There should be no residue at all. Pour through funnel into the volumetric flask and top up to the line so you have exactly 2 L. Store, labeled. Stock solutions B and C are made in the same way, just be careful to accurately measure the small quantities of each ingredient in solution C. All dissolve easily too.

Stock solution D is best made up in a smaller amount, in a 1 L Schott bottle. Start by weighing 5g of sodium hydroxide pellets and work quickly as they absorb water once exposed to air. Dissolve by shaking in 800ml dH₂O for several minutes. It will completely dissolve, but takes a while. Once it has all dissolved add the 32.2g of EDTA and shake again. (See hazards below) Once the EDTA has dissolved too, add the 24ml of ferric chloride solution (60% w/v) and again take care not to ingest any. Once mixed/shaken in, top up to the 1 L line on the Schott bottle. Label solution with its date of preparation.

To make up the dilute solutions for the hydroponic experiments: If you need 1 L, add about 900 mL of water (tap, deionized or distilled) then add the 8 mL of the stocks one by one with stirring in between. This is necessary to ensure that high concentrations of Ca and PO₄do not mix.

Notes and troubleshooting tips

It is convenient and cheap to use tap water for the diluted solutions. The pH of the tap water should be checked first, as the pH of the solutions rise over time as nutrients (especially NO₃) are taken up. Tap water with a high pH (eg high concentrations of Ca carbonates) will need adjusting over time with HCl.

Literature references

Hoagland DR, Arnon DI. 1938. The water-culture method for growing plants without soil. Circular 347, University of California, College of Agriculture, Berkeley.

Health, safety & hazardous waste disposal considerations

• Dry EDTA is a dangerous substance and a face mask and gloves should be worn while dealing with it in its dry form.