Osmolality Explained. Definitions

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1 Osmolality Explained What is osmolality? Simply put, osmolality is a measurement of the total number of solutes in a liquid solution expressed in osmoles of solute particles per kilogram of solvent. When any solute is dissolved in a solvent, four of the colligative properties of the solution are affected in a roughly linear response to the concentration of solute added: The freezing point of the solution is depressed The boiling point of the solution is elevated The osmotic pressure of the solution is increased The vapor pressure of the solution is depressed The resulting changes in these properties are not proportional to the weight, size or shape of the dissolved particles, but only to their molal concentration. Osmolality therefore is an ideal measurement to estimate the total concentration of solutes in a near limitless variety of liquid sample matrices, including blood, serum, plasma, urine, milk, cell culture media and almost all forms of aqueous based solutions. Definitions The following are key terms relevant to the science of osmolality and will help you to develop a better understanding. Avogadro s Number: Number of molecules in one mole (gram molecular weight) of a substance. One mole of non-ionic solute (such as sucrose) dissolved in one kilogram of water will yield Avogadro s number (6.02 x10 23 ) of molecules. One mole of ionic solute dissolved in one kilogram of water will yield almost twice Avogadro s number of particles. Colligative Properties: Properties of solutions that depend on the number of particles in a given volume of solvent, not on the mass of the particles. Colligative properties include: vapor pressure, boiling point, freezing point and osmotic pressure (see also concentrative properties). Concentration: Relative amount of solute in a solution. This can be expressed in many ways: solute to solvent, solute to solution, mass to mass, mass to volume, etc. Concentrative Properties: When a solute is dissolved in solvent, certain properties of the solvent freezing point, boiling point, vapor pressure and osmotic pressure are changed nearly in proportion to the concentration of the solute, expressed in dissolved particles. Avogadro s number of particles, regardless of their size or shape, when dissolved in a kilogram of water, will change each of the concentrative properties a specific amount.

2 Freezing Point Osmometers: Determine the osmotic strength of solution by using freezing point depression. Freezing Point Depression: Describes the phenomenon that the freezing point of a liquid (a solvent) is depressed when another compound is added, meaning that a solution has a lower freezing point than a pure solvent. Ionic Solution: Certain molecules, when dissolved, dissociate into charged particles called ions. A good example is sodium chloride, which dissociates in solution to sodium ions and chloride ions. Membrane Osmometers: Measure the osmotic pressure of a solution separated by a semipermeable membrane. Molality: Molal concentration grams of solute per kilogram of solvent. Molarity: Molar concentration grams of solute per liter of solution. Molecular Weight: The sum of the atomic weights of all the atoms in a molecule. Mole: Gram molecular weight, molecular weight expressed in grams. Each mole contains Avogadro s number (6.02 x1023) of molecules. One mole of sodium chloride weighs grams. Non-Ionic Solution: Certain molecules, when dissolved, do not dissociate or ionize into charged particles. Good examples are glucose and urea. Osmol: Standard unit of osmotic pressure based on a one molal concentration of an ion in a solution. Osmolarity: Osmoles of solute per liter of solution (temperature dependant). Osmolality: Osmoles of solute per kilogram of solvent. Osmotic Pressure: Hydrostatic pressure produced by a difference in concentration between solutions on the two sides of a surface such as a semi-permeable membrane. Solutions: Homogeneous mixture of solutes in a solvent. Solvent: Major, liquid component of a solution. Solutes: Minor components of a solution usually solids. Vapor Pressure Osmometers: Determine the concentration of osmotically active particles that reduce the vapor pressure of the solution.

3 Osmolality Equation Osmolality Equation Osmolality is the number of Osmols of solute particles per kilogram of pure solvent. Since most ionic species do not completely dissociate, osmolality is a unit of concentration, which takes into account the dissociative effect. Osmolality is usually expressed in mosm/kg H 2 0. One milliosmol (mosm) is 10-3 osmols. The osmolality equation is: Osmolality = ΦnC = osmol kg H 2 0 Where: Φ = osmotic coefficient, which accounts for the degree of molecular dissociation n = number of particles into which a particle can dissociate C = molal concentration of the solution Types of Osmometers Types of osmometers An osmometer is a device for measuring the osmotic strength of a solution, colloid or compound. There are three major types of osmometers commerically available, each leveraging a particular colligative property to achieve their analytical results: Freezing Point Osmometers determine the osmotic strength of solution by utilizing freezing point depression Vapor Pressure Osmometers determine the concentration of osmotically active particles that reduce the vapor pressure of the solution Membrane Osmometers measure the osmotic pressure of a solution separated by a semipermeable membrane Advantages and disadvantages of the differerent osmometer technologies are explained Osmometer Type: Advantages : Disadvantages: Comments: Freezing Point Osmometry (FPO) Performs rapid and inexpensive Samples must be of low viscosity FPO provides rapid and inexpensive results with measurements Not ideally suited the industry preferred

4 Simple and reliable performance Industry preferred FP method Small sample size (nl to µl range) Ideal for dilute biological and aqueous solutions for high molality or colloidal solutions freezing point method. Requires small sample size and ideally suited for most biological and aqueous applications. Vapor Pressure Osmometry (VPO) Membrane Osmometry (MO) Performs rapid and inexpensive measurements Small sample size (nl to µl range) Ideal for dilute biological and aqueous solutions Provides potentially unlimited direct measurement of osmotic pressure and solution osmolality Good for colloidal solutions No limitation on sample concentration Can determine MW of macromolecules Less accurate than FPO Cannot be used for volatile solutes like alcohols or other organic solvents Not ideally for high molality or colloidal solutions Time consuming and difficult to operate Requires large sample volume Not applicable for small molecules and aggressive solvents due to membrane porosity and compatibility Irreproducible results due to clogging of membrane pores VPO provides fast an inexpensive measurements requiring a small sample size, although not as fast or reliable as FPO. Volatile solutes are not amenable to VPO limiting its utility in many applications MO provides a direct measurement of osmolality and is suitable for high molality and colloidal samples. Long analysis time and requires a large sample volume. Not applicable for small molecule applications.

5 Osmolarity vs. Osmolality Difference between Osmolarity and Osmolality Measurements of osmolar concentration are often expressed in osmolarity and osmolality. Osmolarity is a measure of the osmoles of solute per liter of solution. A capital letter M is used to abbreviate units of mol/l. Since the volume of solution changes with the amount of solute added as well as with changes in temperature and pressure, osmolarity is difficult to determine. Osmolality is a measure of the moles (or osmoles) of solute per kilogram of solvent expressed as (mol/kg, molal, or m). Since the amount of solvent will remain constant regardless of changes in temperature and pressure, osmolality is easier to evaluate and is more commonly used, and often preferred, in practical osmometry. Most commercially available osmometers report results using osmolality units mosm/kg. As a general rule, a 1 molar solution will have a higher solute concentation than a 1 molal solution. The reason for this is that the solute takes up some volume in the total solution. Consequently, the freezing point depression of a 1 M solution will be lower when compared to the same 1 molal solution. The difference between the freezing point determination will be more noticeable at higher concentrations (i.e 2M, 3M, 4M) and will be alsmost negligable at more dilute concentrations, say 0.5 M or lower. Advantages of Freezing Point Depression Advantages of Freezing Point Depression Freezing point depression osmometers are the industry-preferred solution and the gold standard in clinical chemistry labs, pharmaceutical research, and quality control labs around the world. There are a number of logical reasons for this: It has all of the advantages and none of the disadvantages of other forms of osmometry Membrane osmometry is limited by the lack of a perfect semi-permeable membrane; and vapor pressure osmometry is affected by volatile compounds exerting their own vapor pressure a major drawback in the clinical chemistry lab setting. Its freezing point measurements are fast, accurate and robust Test times are typically less than one minute and sample repeatability is in the range of ± 2 mosm/kg H 2 0. It has small sample size requirements Typical sample volumes are in the nl to µl range, this is advantageous for sample-limited and clinical lab settings. It is simple to use Requires very little knowledge or scientific background to operate systems and achieve good results.

6 It has limitless applicability and utility Freezing point osmometers work well with almost any liquid matrix and are found in nearly every laboratory where osmolality testing is required. It is the most widely referenced and practiced technique for osmolality testing Freezing point osmometers have been commercially available for over 50 years and are the most widely referenced technology for osmolality testing. Properties of Solutions Properties of solutions When solutes are introduced in a solvent, the resulting solution differs from the initial solvent in several ways. The presence of one or more solutes alters the ability of the solvent molecules to interact and reduces their freedom of movement. This affects the solvent s ability to move from solid to liquid or from liquid to gas. These changes are collectively referred to as colligative properties and are dependent on the total number of particles present in the solution. The four basic colligative effects due to the presence of one mole of solute per kg solvent are described in the following : For a simple chemical, such as urea, the effect is related to the total number of moles of urea in solution. For a chemical compound which can dissociate, such as sodium chloride, both the sodium and the chloride ions will contribute to these colligative properties. Therefore one mole of sodium chloride will have twice the effect as will one mole of urea. Using freezing point depression as an example: One mole of urea will depress the freezing point of the solution by 1.86 C One mole of sodium chloride NaCl (1mole Na mole Cl - ) will depress the freezing point by 3.72 C

7 One mole of calcium chloride CaCl 2 (1 mole Ca mole Cl - ) will depress the freezing point by 5.58 C The actual mass of the particle is irrelevant, since a small molecule will exert the same effect as a large molecule.