Cheese making/Experiment 2

Date and Location

Time: 9 pm on Monday July 28, 2015

Location: Omni Oakland, CA.

Random Notes (will be formatted later)

(also see Patrick's email on morning of July 29)

Concern: The accuracy of our pH systems is poor (low-cost eye-dropper/shaker system with range of 6.0 to 7.5, meant for small fishtanks).

Concern: The "5mL" test tubes did not have a marking for the 5mL level (not precise). 

Distilled water: 22.5degC, pH of <6.5 Room temp: 25.2degC

Source of casein: BodyTech 100% micellar casein.(assumed to be acid-casein) Micellar casein, protease (Aminogen®).Contains Milk. 

37.5g of above powder slowly poured into 250mL distilled water while blending at lowest setting for approximately 20seconds. After blending, mixture is strained repeatedly to remove air. Strainer is ~16grid per inch.

Above mixture poured into 5ml test tubes and left to settle for 45 minutes. The pH of this mixture is 6.

After 45 minutes above test tubes placed into D.Poole's sonicator (not the Soni-cat) and sonicated for 30 minutes.  

Prepared a solution of citric acid  2.5g dissolved in 200mL of water. 

Prepared a solution of 5g of baking soda (Nice! brand from Walgreens) in 200mL of water.

Trial and error experiments begin to determine a ratio of the above two solutions that when mixed in 5mL casein/water test tubes result in a slightly alkaline solution of ~7.5pH.  The added citric acid and baking soda solutions varied between 0, 1/8, 2/8, 3/8 of a teaspoon per 5mL test tube. Note, some of the casein/water mixture was poured out to accommodate added baking soda and/or citric acid solutions.

From Patrik's email (pictures still need to be added)

Purpose Test casein powder’s ability to dissolve under different conditions

Methods

12 tubes total; half of the tubes (6) sonicated in jewelry cleaner for 30 min; the others not

Within each set of 6 tubes, half were treated with baking soda (0, 1 or 2 scoops of 1/8 tsp baking soda solution). Th e other half with citric acid then with baking (control; 1 scoop citric acid solution to ~pH6; 1 scoop citric acid + 2 scoops baking soda solution to ~pH7)

all tubes left to settle in fridge overnight

setup - sonicated in blue; baking soda only in front:

Results (see email for pics, which didn’t copy over)

2 scoops of baking soda solution, with or without sonication. The unsonicated casein on the right clearly settled out of solution overnight; the sonicated on the left is still nicely in solution:

Unsonicated, baking soda vs citric acid + baking soda. Not much difference:

Sonicated controls - some settling, but not too much, even without any additions:

Sonicated; baking soda only vs citric acid + baking soda: baking soda only seemed to be marginally better looking at it in person; hard to see on the pics:

Unsonicated controls; this is what happens when you just try to dissolve casein powder in water and let it sit for an hour or two:

Discussion

Sonication can break molecular bonds in polymers; breaks chains and the lower mol wt material is more soluble

    relevance: high power sonication breaks up the larger casein casein powder particles but does not break up the micelle structure itself (paper; Rachel)

Goal: produce recombinant casein, which will not be in powder form

    try and use its change in solubility with pH and Ca levels to make protein crash out of solution for easy prep

Possible future methods:

    3-4 insoluble pellets of different casein proteins
    resolubilize pellets via pH change and Ca concentration
    mix under sonication to let casein micelles form

    use suspended casein powder vs dissolved casein micelles to attempt measuring particle sizes and micelle properties
    control: purified non-micellar casein