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Lipids are one of the major components in food. Some properties of lipids are soluble in organic solvents, non polar (e.g. triacylglycerol and cholesterol) or polar (e.g. phospholipids), and may be present as solid (fats) or liquid (oil) at ambient temperatures. The monomer of lipids is fatty acid that contains an aliphatic chain with a carboxylic acid group. In nature, most fatty acids consist of even number of carbons in a straight chain and vary from 14 to 24 carbons. Short-chain fatty acids have less than 14 carbons and can be found in tropical oils and dairy fats (McClements & Decker, 2008). Fatty acids are also classified as saturated or unsaturated (containing double bonds).
The flavor of food is largely determined by the presence of volatile compounds in the edible fats or oils as the product of lipid oxidation or natural impurities. The perceived aroma and taste of food are often influenced by the type and concentration of lipids present (McClements & Decker, 2008). Free fatty acids are usually related to the development of unwanted properties in food because they can cause off-flavor, reduce oxidative stability, cause foaming, and reduce smoke point. However, the presence of short-chain free fatty acids is sometimes desirable in cheese products where they contribute to flavor profiles.
The two types of lipid chemical deterioration are hydrolytic and oxidative reactions. The production of volatile short-chain free fatty acids that form off-flavor from triacylglycerols is known as hydrolytic rancidity. We use lipase enzyme to study how the enzyme causes hydrolysis rancidity in milk and how the aromas develop in the end product. Interaction of lipids with oxygen or other free radical sources causes a series of chemical reactions that are known as oxidative reaction. During lipid oxidation, the fatty acids esterified to lipids will break down to form small, volatile molecules that produce the off-aromas (McClements & Decker, 2008).
The volatile compounds produced from lipid degradation are generally detrimental to food quality, although these aroma-contributing compounds are preferable in some foods. By understanding the mechanism of lipid oxidation and hydrolytic reactions, we can develop techniques to prevent undesirable lipid degradation. Antioxidants have been used in food industries to prevent lipid oxidation in the food products. These antioxidants have different mechanism in preventing lipid oxidation.
The objectives of the experiment were to distinguish the odors of short chain fatty acids (C2 to C6), identify the enzyme catalyzed rancidity in milk, and study the mechanism of some prooxidants and antioxidants in lipid oxidation.
MATERIALS AND METHODS
The materials and methods used for each lab exercise were listed as follow.
Experiment 1. Odors of free fatty acids.
Some short chain fatty acids (C2 to C6) involved in this experiment were acetic (C2), butyric (C4), valeric (C5), caproic (C6), caprylic (C8), and linoleic (C18:2). The short chain fatty acids were diluted to 500 ppm in water and the odors were carefully smelled and noted.
Experiment 2. Enzyme catalyzed hydrolytic rancidity in milk.
Whole milk and lipase were used in this experiment. Lipase (0.5 g) was added to 100 ml of whole milk and let sit at room temperature for 1-2 hours. Any odors which had developed were noted.
Experiment 3. Lipid oxidation.
For lipid oxidation experiment, the materials needed were phosphotidylcholine liposomes as a source of lipid, buffer pH 7.0, ferric chloride as a source of metal, ascorbic acid, EDTA, propyl gallate, AAPH, thiobarbituric acid (TBA), and water. In 7 separate test tubes, 1 ml of phophotidylcholine liposomes was mixed to 4 ml of buffer pH 7. To each sample, the following catalysts were added:
0.25 ml Ferric chloride (750 ÂÂµM)
0.25 ml Ferric chloride and 0.25 ml ascorbic acid (2.5 mM)
0.25 ml Ferric chloride, 0.25 ml ascorbic acid, and 0.25 ml EDTA (2.5 mM)*
0.25 ml Ferric chloride, 0.25 ml ascorbic acid, and 0.25 ml propyl gallate (5 mM)*
0.25 ml AAPH (50 mM)
0.25 ml AAPH and 0.25 ml EDTA*
0.25 ml AAPH and 0.25 ml propyl gallate*
(* antioxidants were added before prooxidants)
All volumes were brought up to 6 ml with buffer. The solution was incubated at 37 ÂÂ°C for 30 minutes and shook occasionally to mix oxygen in the samples. After 30 minutes, 1 ml of sample was mixed with 2 ml of TBA and was placed in a boiling water bath for 15 minutes. The samples were centrifuged and the absorbance was read at 534 nm.
Table 1. Odors of short chain fatty acids.
Free fatty acids
Strong/sharp, white vinegar scent
Pungent, vomit scent
Strong, scent like aged cheese
Strong, scent like vinegar
Strong, scent like blue cheese, sweet aroma
Very strong scent (resembles body odor)
Figure 1. Thiobarbituric Acid Reactive Substances (TBARS) measurement data on lipid oxidation by several catalysts (ascorbic acid / AA, ethylene diamine tetraacetic acid / EDTA, propyl gallayte / PG, amidinopropane dihydrochloride / AAPH).
1. How does lipase cause rancidity?
1. Describe how do both the prooxidants and antioxidants inhibit lipid oxidation.
2. Do these antioxidants inhibit both prooxidants? Why or why not?
The answers of the following questions have been described in the Discussion.
How does lipase cause rancidity?
Describe how do both the prooxidants and antioxidants inhibit lipid oxidation.
Do these antioxidants inhibit both prooxidants? Why or why not?