- by paper chromatography (qualitative)
- by thin layer chromatography (TLC; qualitative)
- by enzymatic reaction
- by high performance liquid chromatography (HPLC)
- References and further reading
This document gives a brief summary of the procedures and equipment requirements for some commonly used techniques for determination of malic acid in wines. There are several techniques of varying complexity that are used to measure the concentration of malic acid in wine.
Paper chromatography – qualitative
Description: This method is generally used to indicate the presence or absence of an acid. This can assist to monitor the disappearance of malic acid and the presence of lactic acid arising from the malolactic fermentation in wine. Samples of wine and standards are spotted along a line near one edge of the chromatography paper which is then placed in a solvent containing an indicator dye. The distances that the visualised spots move are compared with the standards, to provide identification of the compounds. (Note that the solvents used in this method will require the use of a fume cupboard or other well ventilated work area in compliance with any relevant occupational health and safety requirements.)
Equipment: Flasks, separating funnel, filter papers, capillary tubes, chromatography jar with lid
Reagents: Solvent (requires butanol, formic acid, water, and bromocresol green indicator)
Services: Wash up area, fume cupboard or other appropriately ventilated work area
Space required: Minimal bench space
Thin layer chromatography (TLC) – qualitative
Description: This method is very similar to the method using paper, but has several advantages including being much faster (see Iland et al. 2000). (Note that the solvents used in this method will require the use of a fume cupboard or other well ventilated work area in compliance with any relevant occupational health and safety requirements.)
Equipment: TLC sheets, flasks, capillary tubes, chromatography jar with lid
Reagents: Solvent (requires glacial acetic acid, isobutyl acetate, water), bromocresol green indicator solution
Services: Wash up area, fume cupboard or other appropriately ventilated work area
Space required: Minimal bench space
Enzymatic
Description: The conversion of malic acid by a specific enzyme can be monitored directly by measuring the absorbance (344 nm) resulting from the generation of a by-product of the reaction (NADH). The test is quite straightforward to conduct and requires only sample dilution. Kits for this assay are commercially available.
Equipment: UV spectrophotometer and cuvettes, autopipettes
Calibration: Made-up and kit-supplied standard solutions of glucose in water
Services: Electricity, wash up area
Space required: Bench space depending on spectrophotometer footprint
High performance liquid chromatography (HPLC)
Description: High performance liquid chromatography (HPLC) is a technique that offers several advantages for the analysis of compounds (including malic acid in wine), including potential for automation, high precision and it is relatively fast. However, there is a fairly high capital cost and skilled personnel are required to operate and maintain the instrument. Generally, this technique is only used where there are other applications of HPLC being used by the winery on a routine basis (e.g. organic acids).
Equipment: High performance liquid chromatograph
Calibration: Use made-up standard solutions of malic acid in water
Reagents: Mobile phase, standard solutions for calibration
Services: Electricity
Space required: Significant bench area
References and further reading
- Amerine, M.A.; Ough, C.S. (1980) Methods for analysis of musts and wines. New York Wiley-Interscience.
- Buick, D.; Holdstock, M. (2003) The relationship between acetic acid and volatile acidity. Tech. Rev. (143) 39-43. The Australian Wine Research Institute. Adelaide, SA.
- Iland, P.; Ewart, A.; Sitters, J.; Markides, A.; Bruer, N. (2000) Techniques for chemical analysis and quality monitoring during winemaking. Campbelltown, SA Patrick Iland Wine Promotions.
- Rankine, B.C. (1998) Making good wine: a manual of winemaking practice for Australia and New Zealand. South Melbourne, Sun Books (Macmillan Australia).
- Zoecklein, B.W.; Fugelsang, K.C.; Gump, B.H.; Nury, F.S. (1995) Wine analysis and production. New York Chapman & Hall.