Ashoka Institute Of Technology &

Management
Varanasi

Assignment-

Topic- Testing of neutraceuticals and food products

Submitted by-
Avinash Shukla
B.Pharm 4th Year
 Testing of microbial load-The contaminated food can lead to food poisoning and to ensure
the food safety, the analysis of food is extremely important. Food contamination occurs due to
viruses, bacteria, metals or chemicals. Food poisoning is generally caused due to bacterial
contamination.

The classes of Bacteria associated with food poisoning are E coli, Staphylococci, Salmonella,
Clostridium, Campylobacter, Salmonella and Shigella. They can cause symptoms like nausea,
vomiting, distress, diarrhea, dehydration, and also death. Hence, microbiological testing of food is
important.

Every year, millions of people fall sick and die due to food poisoning. An outbreak of food
poisoning is an alarm for improvement in food safety standards. Regulatory bodies assure the food
processing standards at production, processing, packaging and storage stages and thus ensure
public health. This helps the consumers to avoid food borne illnesses.

Bioburden test-Bioburden is defined as the number of bacteria living on a surface that has not
been sterilized.The aim of bioburden testing is to measure the total number of viable
microorganisms on a medical device, and then use the number to determine the most appropriate
parameters for its final sterilization. Before sterilization, we should perform the test to ensure the
safety and effectiveness of the EO sterilization process.

Total aerobic microbial count-This test determines the total number of aerobic bacteria per mL
or gram of the personal care, cosmetic or pharmaceutical products according USP <61>. This test
gives the total count of bacteria present and requires 48 to 72 hours incubation.The TAMC is a
mandatory test for every lot manufactured of health, beauty, and personal care products .

Total combined yeast and molds count-This test uses aspergillus brasiliensis (ATCC 16404) and
candida albicans (ATCC 10231) to determine the total number of yeast and fungi per mL or gram
of the personal care, cosmetic or pharmaceutical products. This method requires a minimum of
five days of incubation testing.The TYMC is a mandatory test for every lot manufactured of
health, beauty, and personal care products.

Anti-microbial assay-
1.Cup plate method-
Before perform the microbial assay, sterilized the glass ware, sterile the media, antibiotic solution
(standard) and plant samples were kept ready.
The first step of assay involves the preparation of inoculated plates. For this 2% of microbial
suspension is added to the quantity of medium per plate i.e., 0.5 ml of suspension per 25 ml of
nutrient agar medium.Allow the petriplates for solidification for about 10 min.By using sterile
glass (Pyrex) bores, cups were made by maintaining approximate distance between cups (cup
diameter: 6X8 mm2).
Cups were labeled properly to enable the introduction of the test sample, standard and control
precisely. The concerned samples were introduced into appropriate wells with the help of
micropipette; all the cups were filled with equal volumes of sample. To minimize the effect of
variants the petri plates were allowed to store at room temperature for 1-4 hrs, and then the plate
were allowed to incubate for a time period of 1824 hrs.The zone of inhibition was examined and
measured with the help of antibiotic zone reader75-78.

2.Disc Diffusion Method- A geometric series of dilution is prepared for the antibiotic under test
and for the standard preparation. Plates are seeded with the test organism and the medium is
allowed to set on a perfectly horizontal surface so that the agar is constant in depth throughout
the petridish. The organism may be streaked over the medium after it has set. The plates may be
petridish or large flat dishes up to 0.5m.The solution contained in filter paper orcellulose discs,
absorbs a fixed volume of solution. The test and standard solutions are placed in the containers
in random order, to prevent the bias that could be caused by a regular order of plating. The
volume is critical for the cup method but is not significant when cylinders with the lid of the
plate. The plates are left at room temperature for two hours to allow diffusion of the antibiotic to
get a head of growth of the organisms. Then they are incubated at the appropriate temperature,
usually for about 12-16 hours.After incubation, inhibition of growth can be seen as a clear zone
around each container. The diameter of this is proportional to the log concentration of antibiotic.
As soon as possible each diameter is measured byusing an optical system that projects an image
of the plate on to a large grid. Two diameters at right angles are used as a check on ability of the
zone.A graph is plotted for log concentration of standard against zone diameter and results for
the test preparation are plotted on the same graph. Provided the two lines are parallel, the relative
potencies of the standard and the test are represented by the horizontal distance between the two
lines. The parallelism between the two lines can be confirmed mathematically and the potency of
the test obtained by calculation.
Testing of nutritional value- The nutritional value of food refers to the quantity and quality of
nutrients found in the food item, according to the Healthy-food-site.com. Information about the
energy (measured in calories), the macronutrients (carbohydrates, protein, fats), micronutrients
(vitamins and minerals) and phytochemicals of the food are required to understand this.

Laboratory based nutritional analysis- Laboratory based nutritional analysis can be used for
virtually any food type. This method
can be used when:
General Nutritional Labeling needs.
Validating manufacturer / supplier nutritional information.
Validating data generated from software based database analysis.
Product contains unique or exotic ingredients or processing techniques
Analyzing food for animals (pet foods and agricultural feeds).
Accurate nutrient per 100g values are needed for CofAs or Database inclusion.
Reports will feature values per 100g and per serving size (when provided) of the product.
Where applicable, % RDI and % DV values will also be provided. Additional voluntary
nutrients, vitamins and minerals are available please inquire us.

Laboratory Based Nutritional Analysis for Mandatory Nutrients Include:

Protein
Moisture
Ash
Fat Profile (total, sat, mono, poly and trans fat)
Sugar profile
Total Dietary Fiber
Sodium
Calcium
Iron
Cholesterol
Vitamin A
Vitamin C
Carbohydrates
Calories
Calories from Fat
Testing of heavy metals- A heavy metals panel is a group of tests that measures the quantity of
specific potentially toxic metals in the blood, urine or, more rarely, in the hair or other body
tissue or fluid. A laboratory may offer several different groupings of heavy metals panels as well
as tests for individual metals.

Heavy Metals Contamination of Vegetables-

Heavy metal contamination of vegetables cannot be underestimated as these foodstufs are
important components of human diet. Vegetables are rich sources of vitamins, minerals, and
fibers, and also have beneficial antioxidative effects. However, intake of heavy meta-
contaminated vegetables may pose a risk to the human health. Heavy metal contamination of
food is one of the most important aspects of food quality assurance. Heavy metals are no-
biodegradable and persistent environmental contaminants, which may be deposited on the
surfaces and then absorbed into the tissues of vegetables. Monitoring and assessment of heavy
metals concentrations in the vegetables from the market sites have been carried out in some
developed and developing countries.

Heavy Metals/Metalloids-
Any metal (or metalloid) species may be considered a contaminant if it occurs where it is
unwanted, or in a form or concentration that causes a detrimental human or environmental
effect. Metals/metalloids include lead (Pb), cadmium (Cd), mercury (Hg), arsenic (As),
chromium (Cr), copper (Cu), selenium (Se), nickel (Ni), silver (Ag), and zinc (Zn). Other less
common metallic contaminants include aluminium (Al), cesium (Cs), cobalt (Co), manganese
(Mn), molybdenum (Mo), strontium (Sr), and uranium (U).

Calorific Value-
Calorific Value can be performed on a variety of fuels including biomass, oils, coals and
wood.
A weighed quantity of the sample is burned in oxygen in a bomb calorimeter under controlled
conditions. The heat of combustion is calculated from the weight of the sample and the rise in
temperature. The value obtained is the Gross heat of combustion at constant volume,
expressed as MJ/Kg.

Neutraceutical label claim testing- One of the key ways of ensuring label claim compliance
and overall quality of your product is rigorous testing. At Summit Nutritional Laboratories, we
offer testing for most vitamins, botanicals, minerals, amino acids, enzymes, and dietary
supplements. Our experienced and competent staff makes testing your product their top
priority.With the current GMP standards, quality testing is an indispensible part of the
 manufacturing process. Testing is required to ensure that nutraceuticals, nutritional
supplements, and dietary products meet their label claims. HPLC and ICP-MS effectively and
accurately measure vitamins, minerals, and other additives present in raw materials and
finished goods.At Summit Nutritional, we offer NIR Identity, TLC, HPLC and ICP-MS
testing for
Vitamins
Minerals
Herbal raw ingredients

1.Agmark certification-
Agmark is a Quality Certification Mark of the Government of India given to products
confirming to scientificallylaid down quality standards.Products commonly certified under
AGMARK-Honey-Ghee, Butter-Chillies, Turmeric, Curry powder, Garam Masalas,Cardamom,
Cumin, Coriander and other curry powders.-Vegetable Oils-Spices-Wheat, Atta, Maida, Suji,
Besan, Pulses and many more items required by customers.

Certificate of Authorisation-
It is given to the packers authorising them tograde their commodity under AGMARK. Period of
Validity will be 5 years.For granting new certificate of authorisation separate application for
each commodity should be produced.
Documents to be given along with application
1.Sketch of the premises
2.Declaration regarding i.Proprietorship/Partnership etc
ii.Ownership of the premises
iii.Ownership of trade brand label
iv.Use of good grade quality containers for packing commodities.

All declarations has to given in five rupees stamp label attested by notary public.
3.A copy of licence from Panchayat/Municipality.
4.Bank reference:-Letter from the bank regarding the transaction to the packer with the bank.
5.List of machineries.
6.Specimen signature of authorised persons attested by the proprietor/managing partner.
7.Medical fitness certificate of employee.

Method of Marking-
(1) A grade designation mark shall be securely affixed to or printed on
each package in a manner approved by the Agricultural Marketing Adviser or
an officer authoized by him in this behalf in accordance with Rule 11of the General Grading and
Marking Rules, 1988;
(2) In addition to the grade designation mark, following particulars shall be Clearly and
indelibly marked on each package:-
(a)Name and address of the packer;
(b)Place of packing/manufacturing;
(c)Date of packing;
(d)Lot/batch number;
(e)Grade;
(f)Seasonof harvest (incase of chilies only);
(g)Net weight;
(h)Maximum retail Price;
(i)Best before---------month------------year.
(3) The inkused for markingon packages shall be of such quality whichmay not contaminate the
product;5-
(4) The authorized packer, may, after obtaining prior approval of the Agricultural Mar
ketingAdviser or anofficer authorizedby him inthis behalf, mark his private trade mark
or trade brand onthe graded packages provided that the same do not indicate quality o
ther that indicating by the grade designation mark affixed to the graded packages in accorda
nce with these rules.

2.Bureau of indian standards- The Bureau of Indian Standards (BIS), the National Standards
Body of India responsible for formulating Indian Standards was established under The Bureau of
Indian Standards Act, 1986. To protect the interest of consumers, BIS operates a Product
Certification Scheme. Under the scheme, BIS grants licenses to such manufacturers who are
capable of producing goods on continuous basis as per relevant Indian Standards. Testing being
necessary adjunct to product quality evaluation, the need for making available required testing
facilities arises. It is in this context that BIS has established a network of eight laboratories of its
own in the country to cater to testing of samples generated from its Product Certification
Scheme.

Laboratory Advisory Committee (LAC)-

The Laboratory Advisory Committee of the Bureau of Indian Standards is one of the six
statutory advisory committees of the Bureau and advises BIS on policy matters relating to
establishment and operation of its laboratories and other laboratory related matters. LAC
members are Bureau members. The LAC advises BIS on policy matters relating to:

(i) Establishment, operation and expansion of laboratories.

(ii) Testing activities to support research and development programme of the Bureau.
(iii) Participation in National and International Accreditation System for testing and also for
calibration.
(iv) Formulation of guidelines for facilitating the Bureau in recognizing any laboratory in India
or abroad for the purpose of testing and associated activities in accordance with sub-rule (2) of
Rule 10.
(v) Other related laboratory matters.

Testing Charges: The testing charges declared by the laboratory for the relevant Indian
Standards for which lab is recognized, shall be valid for a period of three years. However, in the
event of any revision of the Indian Standard or issuance of an amendment to the Indian Standard,
the revision of testing charges shall be considered and made effective with the prior approval of
the Competent Authority.

Laboratory :
(a) The laboratory shall be manned by regular, qualified and competent personnel.

(b) The lab will have complete in-house test facilities for the relevant Indian Standard. It shall
be fully equipped to test the product strictly in accordance with test methods given in the
relevant Indian Standards with exemptions such as:

Specialized test for which limited facility is available in the country or the test which are not
routinely required, e.g. radioactive test in packaged drinking Water and pull out test in HSD /
TMT bars.

The exceptions will be specifically considered and allowed by the Competent Authority on
merit of the case

(c) In case of breakdown of testing facility, subcontracting may be allowed by BIS till the
equipment is repaired or for a period of six months whichever is less. The sub- contractor shall
be accredited as per IS/ISO/IEC 17025 or ISO/IEC 17025 or a BIS recognized lab for the same
parameter. Under any exceptional circumstances subcontracting may be permitted by BIS on
merit of the individual case.

(d) Availability of adequate electrical power and water supply, where such inputs are required
for testing.

(e) Lab shall give declaration regarding applicable statutory clearances.

(f) The lab shall be independent, without having linkage to any organization or its subsidiary
which is a licensee under BIS Product Certification Scheme. If there is any perceived conflict of
interest due to any reason, recognition will not be granted.

(g) All testing related activities including receipt and preparation of samples and all activities
relating to contract review shall be carried out in a single premise. Hence the lab should have
adequate and proper space as well as infrastructure for storage of samples before and after
testing in the same premise.

(h) The fee as prescribed by BIS from time to time shall have to be paid by the laboratory.

(i) Travelling and stay expenses of the auditors shall be borne by the auditee laboratory as per
entitlement of the auditors.

3.Food safety and standards authority of India-(fssai)- An Act to consolidate the laws
relating to food and to establish the Food Safety and
Standards Authority of India for laying down science based standards for articles of food and
to regulate their manufacture, storage, distribution, sale and import, to ensure availability of
safe and wholesome food for human consumption and for matters connected therewith or
incidental thereto.

(1) This Act may be called the Food Safety and Standards Act, 2006.
(2) It extends to the whole of India.
(3) It shall come into force on such date as the Central Government may, by notification in
the Official Gazette, appoint, and different dates may be appointed for different provisions of
this Act and any reference in any such provision to the commencement of this Act shall be
construed as a reference to the coming into force of that provision.

Establishment of Food Safety and Standards Authority of India.

(1) The Central Government shall, by notification, establish a body to be known as the
Food Safety and Standards Authority of India to exercise the powers conferred on, and to
perform the functions assigned to, it under this Act.
(2) The Food Authority shall be a body corporate by the name aforesaid, having
perpetual succession and a seal with power to acquire, hold and dispose of property, both
movable and immovable, and to contract and shall, by the said name, sue or be sued.
(3) The head office of the Food Authority shall be at Delhi.
(4) The Food Authority may establish its offices at any other place in India.

Packaging and labelling of foods.

(1) No person shall manufacture, distribute, sell or expose for sale or despatch or
deliver to any agent or broker for the purpose of sale, any packaged food products which are
not marked and labelled in the manner as may be specified by regulations:
Provided that the labels shall not contain any statement, claim, design or device which
is false or misleading in any particular concerning the food products contained in the package
or concerning the quantity or the nutritive value implying medicinal or therapeutic claims or
in relation to the place of origin of the said food products.
(2) Every food business operator shall ensure that the labelling and presentation of
food, including their shape, appearance or packaging, the packaging materials used, the
manner in which they are arranged and the setting in which they are displayed, and the
information which is made available about them through whatever medium, does not mislead
consumers.

Liability of the manufacturers, packers, wholesalers, distributors and sellers-

(1) The manufacturer or packer of an article of food shall be liable for such article of
food if it does not meet the requirements of this Act and the rules and regulations made
there under.
(2) The wholesaler or distributor shall be liable under this Act for any article of food
which is
(a) Supplied after the date of its expiry; or
(b) Stored or supplied in violation of the safety instructions of the
manufacturer; or
(c) Unsafe or misbranded; or

(d) Unidentifiable of manufacturer from whom the article of food have been
received; or
(e) Stored or handled or kept in violation of the provisions of this Act, the
rules and regulations made there under; or
(f) received by him with knowledge of being unsafe.
(2) The seller shall be liable under this Act for any article of food which is
(a) sold after the date of its expiry; or
(b) handled or kept in unhygienic conditions; or
(c) misbranded; or
(d) unidentifiable of the manufacturer or the distributors from whom such
articles of food were received; or
(e) received by him with knowledge of being unsafe .
Reference-

http://adamsonlab.com/blog/services/over-the-counter/total-aerobic-microbial-
count/ http://www.tast.com.tw/eng/EO4.php
https://www.google.com/search?q=nutritional+value+of+food+definition&oq=nutr
itional+value+of+foods+defi&gs_l=serp.3.0.0i22i30k1l2.53502.54216.0.56520.4.4
.0.0.0.0.254.495.2-2.2.0....0...1c.1.64.serp..2.2.490.f7otVdLbSD0
https://labtestsonline.org/understanding/analytes/heavy-metals/tab/sample/
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3113373/
http://www.summitnutritional.com/analytical-services/label-claim-testing/
http://agmarknet.nic.in/spices.pdf
www.bis.org.in/lab/lrsIntro.asp
http://www.bis.org.in/lab/lrsIntro.asp
http://www.fssai.gov.in/home/fss-legislation/food-safety-and-standards-act.html