Purification Techniques - MCAT Physical

There are two types of distillation. Simple distillation is used to separate molecules that have very different boiling points. Fractional distillation is used to separate molecules with small differences in boiling points. Fractional distillation is often used if the difference between boiling points is less than . In simple distillation, the vapor is immediately collected in a condenser. On the other hand, fractional distillation allows vapor to condense and revaporize several times. These repeated cycles allow fractional distillation to purify the vapor better than simple distillation.

Distillation is used to separate molecules with different boiling points. Simple distillation is used to separate molecules with vastly different boiling points. Fractional distillation, on the other hand, is a refined form of simple distillation that can be used to separate molecules with similar boiling points. Note that fractional distillation can separate molecules with either different or similar boiling points; therefore, fractional distillation can be used to separate any of the given mixtures.

Rate of distillation is increased when the ability of a substance to become a vapor is increased. Recall that vapor is created when enough heat is applied to the liquid. The temperature at which the liquid becomes vapor is called the boiling point. A liquid turns into a vapor when the vapor pressure (pressure applied by the vapor from the liquid) equals the atmospheric pressure. Decreasing the atmospheric pressure will make it easier for the liquid to turn into a vapor; therefore, this will increase the rate of distillation.

Decreasing the vapor pressure will remove vapor from system. This will make it harder to distill substances. Decreasing temperature will move the system away from the boiling point, thereby decreasing the amount of vapor. Decreasing mole fraction of the substance will decrease the surface area of the substance (at the surface of the solution). Liquid molecules need to be present at the surface to escape the solution and become vapor; therefore, decreasing mole fraction will decrease the amount of vapor.

Gel electrophoresis is a technique used to separate molecules based on size or charge. Charged particles can be separated because they migrate towards different ends of the gel.

Negatively charged particles always migrate towards the positive pole whereas positively charged particles always migrate towards the negative pole (opposites attract). In gel electrophoresis, the positive pole is called the anode and the negative pole is called the cathode; therefore, the charged particles will migrate to the respective nodes.

The isoelectric point is the pH at which a charged particle loses its charge and becomes neutral; therefore, the pI is only a property of the species (not its environment). When the pH of the environment equals the pI of the molecule, the electrical charge on the molecule disappears. A basic environment is characterized by a high pH. If the pH of the environment is higher than the pI, then the environment is basic. The size of the species is usually represented by mass or weight, not by pI.

pI of a substance affects other chemical properties, such as solubility. Charged species are polar and are likely to dissolve in polar solvents whereas uncharged species (when pH = pI) are likely nonpolar and are more soluble in nonpolar solvents.

A pH gradient in a gel facilitates the separation of charged species. The proteins in the mixture encounter different pHs as they travel through the gel. The driving force for the proteins is the electric force. Recall that each protein has an isoelectric point; when the protein's environmental pH equals the isoelectric point, the protein will become neutral and stop migrating (it loses the electric force). The physiological pH is about 7.4. Several amino acids are uncharged at this pH; however, these amino acids will become charged when presented in a different pH environment; therefore, these proteins will be charged and be able to migrate through the gel because of the pH gradient.

The protein will stop migrating when the pH of the environment (gel) equals the pI of the protein. Different types of gel electrophoresis are used to separate molecules based on charge and size. A pH gradient is used to separate molecules based on charge, whereas SDS-PAGE is used to separate molecules based on size. All types of macromolecules (nucleic acids, proteins, lipids, and carbohydrates) can be separated using gel electrophoresis.

SDS is a substance added to polyacrylamide gels to separate substances based on size. This method is called SDS-PAGE. SDS binds to the secondary structure of proteins and gives them an overall negative charge; the bigger the substance the bigger the negative charge. The larger mass compensates for the bigger charge; therefore, SDS makes it so that the mass-to-charge ratio of all substances is the same. This standardizes the electric force experienced by each molecule, making size the only driving force for migration through the gel.

TLC is a laboratory technique commonly used to separate components of a mixture. Mixtures are placed on the TLC plate (stationary phase), which is then transferred to a jar containing the solvent. The solvent travels through the plate and carries components of the mixture along with it. Based on its properties, each component is dragged to different distances on the plate. The relative distances travelled by each component can be used for separation and identification.

It is important to place a lid on the jar because the solvent will be a volatile substance. An open system will allow for the solvent to evaporate from the TLC plate and reduce the amount of solvent travelling through the plate. The solvent in the solution will evaporate, but it is negligible and inconsequential to the data collected on the TLC plate.

Thin layer chromatography (TLC) is used to separate components in a mixture. Components are separated on a TLC plate because each component travels a different distance. The distance travelled depends on several factors. One of those factors is polarity; therefore, TLC can used to determine polarity of substances.

TLC is not useful for identifying substances. Other techniques such as NMR and IR spectroscopy are useful for identifying individual components. The Rf value of a compound can help identify it in a broad sense, but this technique provides only very rudimentary identification capabilities.

To solve this problem we need to recall the definition of Rf.

The solvent is typically the most mobile substance on a TLC plate; therefore, it travels the farthest distance. This means that Rf is always less than one because the numerator of Rf is always smaller than the denominator. The question states that the Rf value is 2. Since this is greater than 1, the results don’t seem valid. This value indicates that the solute compounds were able to teravel farther on the TLC plate than the solvent (mobile phase).

Compounds A and B are in the same mixture; therefore, they will be separated using the same TLC plate. Recall the definition of Rf.

Since the same TLC plate was used, the denominator for both compound A and B will be the same (the total distance travelled by solvent alone is the same for a TLC plate). To separate them effectively, compounds must have travelled different distances along the plate. The best separation occurs when the difference between the distances travelled by the compounds is greatest.

Let’s assume the denominator to be 1 for both compounds. This means that the numerator is equal to the Rf value. If we compare the given Rf values with each other we realize that Rf values of 0.4 and 0.2 will give the biggest difference between the distances travelled by compounds A and B (difference is ); therefore, this will have the best separation.

Distillation is a process of separating a liquid from solutes or other liquids. It utilizes the boiling point differences to separate substances. A substance with a low boiling point will evaporate first in a distillation column and will be isolated first. The question is asking which solution will be isolated first; therefore, we need to figure out which solution has the lowest boiling point. Recall that the boiling point of a solution is elevated when there are more solutes present in the solution. Sodium chloride () contributes two solutes (sodium ions and chloride ions). Calcium nitrate () contributes three solutes (one calcium ion and two nitrate ions). Sucrose does not dissociate into ions in solution; therefore, it only contributes one solute. This means that the sucrose solution will have the lowest amount of molecules in solution, the lowest boiling point, and will be separated first.

There are two types of distillation. Simple distillation is used to separate molecules that have very different boiling points. Fractional distillation is used to separate molecules with small differences in boiling points. Fractional distillation is often used if the difference between boiling points is less than . In simple distillation, the vapor is immediately collected in a condenser. On the other hand, fractional distillation allows vapor to condense and revaporize several times. These repeated cycles allow fractional distillation to purify the vapor better than simple distillation.

Distillation is used to separate molecules with different boiling points. Simple distillation is used to separate molecules with vastly different boiling points. Fractional distillation, on the other hand, is a refined form of simple distillation that can be used to separate molecules with similar boiling points. Note that fractional distillation can separate molecules with either different or similar boiling points; therefore, fractional distillation can be used to separate any of the given mixtures.

Rate of distillation is increased when the ability of a substance to become a vapor is increased. Recall that vapor is created when enough heat is applied to the liquid. The temperature at which the liquid becomes vapor is called the boiling point. A liquid turns into a vapor when the vapor pressure (pressure applied by the vapor from the liquid) equals the atmospheric pressure. Decreasing the atmospheric pressure will make it easier for the liquid to turn into a vapor; therefore, this will increase the rate of distillation.

Decreasing the vapor pressure will remove vapor from system. This will make it harder to distill substances. Decreasing temperature will move the system away from the boiling point, thereby decreasing the amount of vapor. Decreasing mole fraction of the substance will decrease the surface area of the substance (at the surface of the solution). Liquid molecules need to be present at the surface to escape the solution and become vapor; therefore, decreasing mole fraction will decrease the amount of vapor.

Gel electrophoresis is a technique used to separate molecules based on size or charge. Charged particles can be separated because they migrate towards different ends of the gel.

Negatively charged particles always migrate towards the positive pole whereas positively charged particles always migrate towards the negative pole (opposites attract). In gel electrophoresis, the positive pole is called the anode and the negative pole is called the cathode; therefore, the charged particles will migrate to the respective nodes.

The isoelectric point is the pH at which a charged particle loses its charge and becomes neutral; therefore, the pI is only a property of the species (not its environment). When the pH of the environment equals the pI of the molecule, the electrical charge on the molecule disappears. A basic environment is characterized by a high pH. If the pH of the environment is higher than the pI, then the environment is basic. The size of the species is usually represented by mass or weight, not by pI.

pI of a substance affects other chemical properties, such as solubility. Charged species are polar and are likely to dissolve in polar solvents whereas uncharged species (when pH = pI) are likely nonpolar and are more soluble in nonpolar solvents.

A pH gradient in a gel facilitates the separation of charged species. The proteins in the mixture encounter different pHs as they travel through the gel. The driving force for the proteins is the electric force. Recall that each protein has an isoelectric point; when the protein's environmental pH equals the isoelectric point, the protein will become neutral and stop migrating (it loses the electric force). The physiological pH is about 7.4. Several amino acids are uncharged at this pH; however, these amino acids will become charged when presented in a different pH environment; therefore, these proteins will be charged and be able to migrate through the gel because of the pH gradient.

The protein will stop migrating when the pH of the environment (gel) equals the pI of the protein. Different types of gel electrophoresis are used to separate molecules based on charge and size. A pH gradient is used to separate molecules based on charge, whereas SDS-PAGE is used to separate molecules based on size. All types of macromolecules (nucleic acids, proteins, lipids, and carbohydrates) can be separated using gel electrophoresis.

SDS is a substance added to polyacrylamide gels to separate substances based on size. This method is called SDS-PAGE. SDS binds to the secondary structure of proteins and gives them an overall negative charge; the bigger the substance the bigger the negative charge. The larger mass compensates for the bigger charge; therefore, SDS makes it so that the mass-to-charge ratio of all substances is the same. This standardizes the electric force experienced by each molecule, making size the only driving force for migration through the gel.