Keratin is a fibrous protein and has a structural function. Fibrous proteins contain polypeptides that bind together to form very long fibres running parallel to each other. These fibres are linked by cross-bridges that make the protein strong and insoluble.
The main constituent of hair is the protein keratin. Keratin can also be found in horns, nails, claws and feathers. These examples suggest keratin has a strong structure. Like most other proteins it is made of monomers called amino acids. All Amino acids contain both an acidic carboxylic group (-COOH) and a basic amino group (-NH2); there is also an R group that changes with each different amino acid. They all consist of carbon, hydrogen, oxygen and nitrogen atoms. Keratin also contains a high concentration of the amino acid cysteine; this contains a sulphur atom. The sulphur atoms from two cysteines join together, forming a very strong disulphide bond. These bonds are covalent and form strong links making the tertiary structure of the protein very stable. The disulphide bonds occur down the length of the keratin fibre and the cross-linking between the keratin chains account for the strength of hair. Within each hair strand the keratin chains are also linked with ionic, salt and hydrogen bonding. Hydrogen bonding occurs from the attraction between the electronegative oxygen atoms on the CO groups and the electropositive H atoms on either the OH or NH groups. Although they are individually weaker than disulphide bonds, hydrogen bonds are in much higher proportions to the disulphide bonds making them important in maintaining the tertiary structure of the protein.
Hair is very resilient and has elastic properties. These elastic properties are also due to the flexibility of the hydrogen bonds. An untreated healthy hair can support a load of up to approximately 100g. The strong disulphide bonds in the cortex help to give strength to keratin. The long keratin molecules in the cortex are compressed to form a regular structure, which is not only strong but also flexible.
Protein structure and their properties are altered by the change in pH values. Keratin is therefore affected by the pH. The pH of a solution is the measure of how acidic or alkaline it is. An acidic solution (pH<7) has a greater abundance of H+ ions and a basic solution (pH>7) has more OHï¿½ ions present. Most proteins are stable over a limited range of pH values. Outside this range free hydrogen ions or hydroxyl ions affect the charges on amino acids residues, distorting the three-dimensional shape and can cause irreversible change in the protein’s tertiary structure. At very low pH all amino acids exist as ions with an overall positive charge, while at high pH they exist as ions with an overall negative charge.
Protein structure can be easily disrupted by changes in pH value away from the optimum range. Unfolding of the formation due to pH is a result of ionisation of the R group inside the protein molecule. This damages and breaks the ionic and hydrogen bonds that help to hold the protein and give strength. Consequently weakening the hair strand and causing it to lose its elasticity.
The hydroxyl ions can split the disulphide bonds, removing the cross-links between keratin chains, weakening the hair. This is why it is possible for strong alkali solutions to dissolve protein.
Most proteins have a buffer system that can resist changes in pH and excrete excess acid or alkali. An amino acid is amphoteric and means that slight changes in pH will not affect the keratin. Small changes in pH will be resisted and neutralised by the natural buffer in the amino acid. However, when submerged in strong pH solution especially alkaline solutions, the increase in Hydrogen or hydroxyl ions will cause ionisation of the important bonds holding the keratin fibres together. This will cause damage to the bonds in the hair strand leaving it with less resilience and less strength.
To test my prediction I shall immerse individual hair strands in solutions of five (varying) pH values, then apply weights gradually to the individual hairs and record the mass needed to break the hair strand.
A gradual increase in the mass added to the treated hair is essential so that an accurate strength can be determined. If a large force is applied suddenly to the hair, the tensile strength is reduced and the hair has no time to respond and stretch. A steady increase in force will enable the distribution of the pressure over the length of the hair fibre so there is some resistance. This means that the weight applied must begin relatively small then gradually increased.
The first mass that will be applied to the treated strands of hair is 20g then after a thirty second interval five grams will be added so 25g, then 30g, 35g, 40g and continues in this five-gram interval until the hair eventually snaps. This ensures a build up of mass, which is steady and also means that a more accurate measurement can be taken of what mass breaks the hair. The thirty-second period between applying five grams more is to ensure that the hair has stretched.
The alternative method of testing hair strength would be to draw the hair apart with hands, but this method means a difficulty in comparisons and also is very inaccurate.
I will use a sample of a hundred (human, head) hair strands of the same hair type and of equal 20cm lengths. The length of hair needs to be 20cm so that there is a sufficient length to fasten the hook and also so the mass can be more evenly distributed.
A hair strand will be taped to a wooden splint, with one end fastened, and the other free to be immersed into the test solutions. There will be five different splints to be used for the five different pH solutions in test dishes. The five test dishes will be cleaned in pH neutral solution and rinsed with distilled water, this is to prevent contamination. The five pH solutions will have varying values across the pH scale. A strong acid (pH 1-4), a weak acid (pH 4-6.4), a neutral (pH 7), a weak alkali (pH 7.5-9.9) and a strong alkali (pH 10-14) will be used to test individual hairs. This range is representative of the principal pH values. The chemical compounds chosen to represent these pH values are Hydrochloric Acid, Acetic Acid, Water, Calcium Hydroxide and Sodium Hydroxide respectively. These compounds are commonly used in the laboratory and practical to obtain. Also their pH values correspond to the values chosen to test. Each test dish will have a 2cm3 chemical solution with a concentration of 0.5M. This concentrate is the safest amount to use whilst still getting a strong pH.
The five separate experiments will be conducted simultaneously so that all the conditions are the same. The dishes and splints must be labelled accordingly with their pH values. The corresponding splint will be placed into the solution, the hair is to be exposed for 10 minutes, to ensure that the hair has been exposed thoroughly. The hair will be removed gently and rinsed with distilled water and allowed to stand to air dry. The hair is rinsed in order to equilibrate at this pH.
Buffer solutions are not used as they may interfere with the amphoteric nature of the amino acid in keratin. Instead to record the exact acidity or alkalinity of the solutions a pH probe will be used. Indicators such as the pH dyes and paper strips are both inaccurate and are affected by other components in the solution. A pH probe is very accurate and recordings are precise. However before usage the probe must be calibrated and tested. The probe should be cleaned in distilled water before usage, to remove contamination. Each solution should be mixed with a magnetic stirrer, and then the probe should be placed horizontally in the dish, allowing sufficient time for the pH to stabilise. After each reading the probe should be rinsed.
After the hairs have dried a hook will be tied to the free end of the hair and the hair placed in a stand. From this hook weights can be hung. A Vernier Callipers will be placed by the side of the hair fibre to measure the length each hair stretches. This device is accurate to 0.1mm and is suitable for the measurement. I will measure the length of the hairs at different weights, and also record the weight needed to break the hair. This data can be plotted onto a graph to create different curves. This way hair strength can be calculated.
The more repeats of each experiment, the clearer the strength of the hairs immersed at different pH, will become. Statistical analysis (standard deviation) is used to eliminate any variance within treatments and calculate a realistic value. I have decided to carry out each experiment at the five different pH’s fifteen times. This is to ensure accurate results, which are not affected by anomalous data. This many repeats is also necessary because the experiment is very fragile and there are likely to be faults made which affect a few results.
To produce reliable results there are certain variables that must be controlled in each experiment. The humidity, temperature and the hair width must all be kept constant. Temperature affects the dissociation of water, and therefore the pH of a neutral solution changes with temperature. The temperature must be maintained at 20-25ï¿½c. The pH probe must be calibrated at this temperature to ensure correct readings. Thermostatically controlled temperature rooms are used to maintain temperature. A thermometer will then be used to check that the area of experimentation is at the correct temperature.
The humidity of the atmosphere can affect the hair fibre; the hair can absorb moisture, which damages hydrogen bonds and weakens the hair. Keeping the humidity level around the experiment area as constant as possible is essential. The first and simplest precaution taken will be to position the experiment away from areas where it would be exposed to extremes of temperature and humidity such as heating and cooling vents, stoves, doors and windows. A humidifier/de-humidifier would cost too much to install and is impractical. To monitor changes in humidity a hygrometer will be used. Hygrometers are used to measure the relative humidity of air. These are simple to use and are low cost. Direct sunlight is especially damaging, so the position and time of day the experiment is conducted must not alter for the repeats.
The sample hairs should all have approximately the same cross sectional area all the way down; so the hairs will be cuttings from the crown of the head. The hair thickness should be similar from the same area. The cross- sectional area of the hair can influence the strength of the hair, so this must be kept constant (the thicker the hair the added strength).
There are trials that must be conducted before the experiment to ensure apparatus and resources are responding correctly, and to make judgements for the experiment. The pH probe must be tested several times before usage. The probe must be placed in a standard solution of known pH (at a temperature between 20-25ï¿½), if the correct reading appears on the meter then the experiment can commence, if not the probe must be adjusted.
The strength of strands of hair that is untreated must be calculated to make comparisons, and also in order to understand the effect of pH on keratin. The untreated hair strands will also have weights applied to them and the mass needed to break the hair will also be recorded. This test will also be carried out fifteen times, exactly the same as the treated hair fibres.
Hydrochloric Acid – CORROSIVE
May cause burns. The vapour is very irritating to the respiratory system. I am to use a 0.5M, which is considered an IRRITANT. All solutions will be handled with protective gloves on, and protective eyewear must be worn at all times. A laboratory coat is also to be worn. Hair tied up. Keep at a distance from bases.
General Use: Wear eye protection and gloves. Used with careful supervision.
Acetic Acid- CORROSIVE
Causes severe burns; avoid contact with liquid or vapour in the eyes, on the skin, by breathing, or on clothing. Solution to be kept in a cool place and avoid liquid or vapour contact. In the 0.5M solution to be used the chemical is an IRRITANT.
General Use: Gloves must be worn and protective eyewear as a precaution. Use bottle carriers for transporting acid bottles. Segregate acids from bases.
Calcium Hydroxide- CORROSIVE
Inhalation causes irritation to the respiratory tract. Symptoms may include coughing, shortness of breath. Calcium hydroxide is CORROSIVE
General Use: Gloves and eyewear must be worn at all times to reduce exposure to hands.
Sodium Hydroxide- CORROSIVE
Causes severe burns. Very dangerous to eyes and skin. Solution will be equal to 0.5 M and should be labelled CORROSIVE.
General Use: Eye protection is very important. The solution must not be handled directly. The use of beakers with handles is safer, as they are less likely to be dropped
Ethical Implications: The donor of the hair, must consent to the manner the hair will be used and agree with the testing. The hair should be chosen from a healthy individual who does not suffer from a hair disorder.