Analytical tools in Nutrition & Dietetics.

HND-408
Ma’am Sehrish comp. by: Huzaifa 1455

Importance of moisture determination:

Moisture Assay:
Moisture is one of the most fundamental and important analytical procedure that can be
perform on a food product. The dry product that remains after moisture removal is commonly refer as
total solids.
Importance:
• Moisture is a quality factor in the preservation of same products.
• Reduced moisture is used for convenience in packaging or shipping.
Form of water in food:
1. Free water: This water retains the physical properties and act as dispersing agents for colloids
and solvent for salts.
2. Absorbed water: This water is held tightly in cell walls and is held to protein.
3. Water of hydration: This water is bound chemically. e.g. lactose monohydrate, Na2SO4, 10H2O
Practical. Determination of moisture content
Sample preparation.
The objective of preparation of sample is to obtain a representative sample (portion of food that
represent the whole batch)
Only edible portion… From food, vegetables, meat, fish, nuts are taken for sample. These are normally
crushed and minced into fine and homogenous.
On liquid foods… all liquid food such as milk, juices, oil are mixed well before taking the representative
sample. Similarly, cereals, grains, legumes, are ground to produce powder form. Food such as butter are
taken as such for sample.
Procedure:
▪ Weigh an empty clean and dry Petry dish
▪ Place the sample in weighed petry dish.
▪ Place the dish in hot air oven at 100 C for 4 hrs.
▪ Remove petry dish after 4 hrs and cool in desiccator.
▪ Place the dish again in oven for 2 hrs.
▪ Repeat the procedure till constant reading is obtained.
Moisture= initial weight of sample—weight of dry sample X 100
Weight of sample
Introduction to food analysis
Objective
• Govt regulation require for certain products with standers of identity.
• Nutritional labeling regulation require.
• Quality control monitor product Quality for consistency.
• Research and development of new products and improving existing products.
Properties
Chemical properties (fat, protein, carbs, macro/micro nutrients)
Physical properties (stability, shape, hardness)
Sensory properties (taste, flavor, color, texture)
Reference of official methods
• AOAC ( association of analytical chemist)
 • AOCS (American oil chemist society)
• AACC (American association of cereal chemist)
Criteria for selecting analytical technique:
There are many techniques to analyze but each has probes. Select the technique that is require or fit
into your system.
• Precision
• Accuracy
• Simplicity
• Cost
• Sensitivity
• Reproductivity
• Official approval
Sample
• Official sample
• Finish product (prepared)
• Raw sample
Sampling plan is predetermine procedure for selection withdrawal preservation transportation on
preparation of the portion to be removed from a lot as sample. The sampling plan should be clearly
written document containing details. Such as;
• Number of samples selected
• Sample location
• Method of collecting sample
• Factor affecting
Factor affecting sample; purpose of inspection/analysis, nature of product (homogenesis)
Nature of test method:
➢ Nature of population
➢ Sample selection
➢ Sub-sample selection
➢ Sample of selection can be manually a mechanically (as in industries)
Practical part principle:
Oven dry method involved the removal of moisture from sample and the weight determination of solids
remaining to calculate moisture content. Non water volatile can be loss during dry but their loss is
negligible percentage of amount of water loss. Distillation procedure also involve a separation of
moisture from solid and moisture is determined directly by volume.
Karl fisher
Karl fisher titration is based on chemical reaction of moisture present reflected as the amount of titrant.
Di-electric method is based on electrical properties of water.
Nature of sample
• Most of food will Tolerate oven drying at high temperature. Some food contains volatile that are
losed at such temperature (high)
• Some foods here constituents that undergo chemical reactions at high temperature.
• Vacuum over drying at reduced temperature may overcome some such problems.
• for food in moisture or high in fat and sugar Karl fisher titration is best method.
• The use of hydrometer and refractometer used for check sugar liquid samples.
• The free water present in food is more easily determine as compared to bound water.
Fat components
Fat = insoluble in water and soluble in organic solvent such as ether, alcohol, benzene etc.
Types of fat:
Saturated fats (single bond)
Unsaturated fats (double bond)
Classification of fat:
Simple; fatty acid, waxes(fatty+alchol) , oil (fatty acid + glycerol)
Compound: phospholipids, lipoprotein
Derived; (formed by breakdown of simple and compound lipids)
fat soluble vit, simple fatty acid, sterol, alcohols.
Importance of analysis:
An accurate and precise quantitative and qualitative analysis of lipids in foods is important for accurate
nutritional labeling, determination of whether the food meets the standard of identity, and to ensure
that the product meets manufacturing specifications. Inaccuracies in analysis may prove costly for
manufacturers and could result in a product of undesirable quality and functionality. water insolubility is
the essential analytical property used as the basis for the separation of lipids from proteins, water, and
carbohydrates in foods.
Solvent extraction:
The total lipid content of a food is commonly determined by organic solvent extraction methods or by
alkaline or acid hydrolysis. For multicomponent food products, acid hydrolysis is often the method of
choice. Both acid hydrolysis and alkaline hydrolysis methods can be performed before organic solvent
extraction. The use of acid hydrolysis eliminates some of the matrix effects that may be exhibited by
simple solvent extraction methods. The accuracy of direct solvent extraction methods depends on the
solubility of lipid in the solvent used and the ability to separate the lipid from complex from
macronutrients.
Properties of Sample Preparation:
The validity of the fat analysis of a food depends on proper sampling and preservation of the sample
before the analysis. An ideal sample should be as close as possible in all of its intrinsic properties to the
material from which it is taken. The sample preparation for lipid analysis depends on the type of food
and the type and nature of lipids in the food. The extraction method for lipids in liquid milk is generally
different from that for lipids in solid soybeans. For effective analysis of lipid structure chemically class
and components.
Preparation of sample:
Dry the product and remove moisture in order to facilitate the entry of organic solvent because
moisture restrict the entry of organic solvent. Size reduction is done to increase the surface area of
sample after this acid hydrolysis is done which helps in breaking protein, fat emulsion and increase the
availability of fat solvent.
Requirements; weighing balance, soxhled apparatus, drying oven, thimble, heating mentle, glass rod,
desiccator, solvent (hexane, diethyl ether 60-80°C) cotton plugs.
Procedure: (protocol method)
o Weigh 5g of grounded and dried sample and place it in the thimble.
o Take a 150ml round bottom flask, clean and fill the flask 75-100ml solvent.
o Place the whole apparatus on heating metal and allow the diethyl ether to boil.
o Continue the extraction process for 6-7 hrs. separate the condensing unit from extraction unit
and allow the sample to cool.
o Place the thimble with sample in oven after removing the sample, place it in desiccator, weight
the thimble with sample.
FAT= weight of thimble with sample – empty thimble X 100
Weight of sample
Principles of fat Assay
Semicontinuous Solvent Extraction Method: Soxhlet Method
For semicontinuous solvent extraction, the solvent builds up in the extraction chamber for 5–10 min and
completely surrounds the sample and then siphons back to the boiling flask. Fat content is measured by
weight loss of the sample or by weight of the fat removed. This method requires more time than the
continuous method. Instrumentation for a more rapid and automated version of the Soxhlet method is
available.
Non-solvent wet extraction method Gerber Method for Milk Fat (same as Babcock method)
In the Babcock method, H2SO4 is added to a known amount of milk in the Babcock bottle. The sulfuric
acid digests protein, generates heat, and releases the fat. Centrifugation and hot water addition isolate
fat for quantification in the graduated portion of the test bottle. The fat is measured volumetrically, but
the result is expressed as percent fat by weight.
Procedure of Gerber Method, Babcock:
•Transfer 10ml H2SO4 at 15-20°C into a Gerber milk bottle.
• accurately measure milk sample (11ml) into the Gerber bottle using a Gerber pipette.
• add 1ml of iso-amyl alcohol to the bottle
• tighten the stoper and mix by shaking the bottle.
• centrifuge the bottle for 4mint.
• place the bottle in a water bath at 60-63°C for 5mint and then read fat content from the graduation on
bottle neck.
Applications
The Gerber method is comparable to the Babcock method but is simpler and faster and has wider
application to a variety of dairy products. The isoamyl alcohol generally prevents the charring of sugar
found with the regular Babcock method. This test is more popular in Europe than in America.
Ash Analysis
Ash is inorganic residue remaining after ignition or complete oxidation of organic matter in food stuff.
Types
•Dry Ashing
•wet Ashing
Dry Ashing
Dry ashing is used for some type of proximate composition. Dry ashing refers to the use of a
muffle furnace capable of maintaining temperatures of 500–600◦C. Water and volatiles are vaporized,
and organic substances are burned in the presence of oxygen in air to CO2 and oxides of N2. Most
minerals are converted to oxides, sulfates, phosphates, chlorides, and silicates.
Wet Ashing
Wet ashing is used of some minerals. Most of dry sample need no preparation while fresh
vegetables need to be dry before ashing. Wet ashing is a procedure of oxidizing of organic compounds
by using acids. Minerals are solubilized without volatilization. Wet ashing is preferable as compare to dry
ashing.
Importance of Ash in Food Analysis
Ash content represents the total mineral content in foods. Determining the ash content may be
important for several reasons. It is a part of proximate analysis for nutritional evaluation. Ashing is the
first step in preparing a food sample for specific elemental analysis. Because certain foods are high in
particular minerals, ash content becomes important.
Sample Preparation
It cannot be overemphasized that the small sample used for ash, or other determinations, needs to be
very carefully chosen so that it represents the original materials. A 2–10-g sample generally is used for
ash determination. For that purpose, milling, grinding, and the like probably will not alter the ash
content much; however, if this ash is a preparatory step for specific mineral analyses, contamination by
microelements can occur.
Principle of dry ashing
Dry ashing is incineration at high temperature (525◦C or higher). Incineration is done with a muffle
furnace. Crucible selection becomes critical in ashing because the type depends upon the specific use.
Crucible should be mark for the identification.
Advantages
Advantage of dry ash is that it is a safe method. It requires no added reagent but little attention
is needed when ignition begins. Large number of crucibles can be handled at same time and resulted ash
can be used in analysis.
Disadvantage
The disadvantages are the length of time required (12–18 h or overnight) and expensive
equipment. There will be a loss of the volatile elements and interactions between mineral components
and crucibles. Volatile elements at risk of being lost include As, B, Cd, Cr, Cu, Fe, Pb, Hg, Ni, P, V, and Zn.
Protein analysis
Importance of protein analysis.
Protein analysis is important for:
1. Nutrition labeling
2. Pricing: The cost of certain commodities is based on the protein content as measured by nitrogen
content (e.g., cereal grains; milk for making certain dairy products, e.g., cheese).
3. Functional property investigation: Proteins in various types of food have unique food functional
properties: for example, casein in milk for coagulation into cheese products, and egg albumen for
foaming.
4. Biological activity determination:
Protein analysis is required when you want to know:
1. Total protein content
2. Content of a particular protein in a mixture
3. Protein content during isolation and purification of a protein
4. Nonprotein nitrogen 5. Amino acid composition.
6. Nutritive value of a protein
Principle Kjeldahl method
In the Kjeldahl procedure, proteins and other organic food components in a sample are digested with
sulfuric acid in the presence of catalysts. The total organic nitrogen is converted to ammonium sulfate.
The digest is neutralized with alkali and distilled into a boric acid solution. The borate anions formed are
titrated with standardized acid, which is converted to nitrogen in the sample. The result of the analysis
represents the crude protein content of the food.
Applications
Advantages:
1. Applicable to all types of foods
2. Inexpensive (if not using an automated system)
3. Accurate; an official method for crude protein content
4. Has been modified (micro Kjeldahl method) to measure microgram quantities of proteins
Disadvantages:
1. Measures total organic nitrogen, not just protein nitrogen
2. Time consuming (at least 2 h to complete)
3. Poorer precision than the biuret method
4. Corrosive reagent
Proof reading by: Ayesha Aziz, Sadia Parveen, Sumaira Ashraf