The Scientific Panel on the Methods of Sampling and Analysis, Scientific Committee and the Food Authority has approved the methods for the analysis of fortificants in various food products. These can be checked in Annexure I below.
The FSSAI has also requested the food testing laboratories to use these same methods for the analysis of fortificants. If there are any issues related to these methods then these may be forwarded to the Scientific Panel on Methods of Sampling and Analysis for its consideration at the following email: spsampling@fssai.gov.in
Annexure 1
Instrumentation based methods for analysis of Fortificants in Foods
S.N |
Commodity |
Parameter |
Test method |
Brief sample size and extraction process, Mobile phase solvent |
Instrumentation and Detector/ Method |
1. |
Salt |
Iodine content |
AOAC 925.56 (Titrimetric) |
Microwave digestion with nitric acid. pH adjustment with ammonium hydroxide. Alkalization helps to convert volatile iodine to more stable iodide or Iodate. Sodium iodide used as intermediate standard. |
|
|
|
Iron content |
AOAC 2011.14 (ICP) AOAC 985.35 and AOAC 999.10 (AAS) AOAC 2015.06 (ICP-MS) |
Microwave assisted Inductively Coupled Plasma Optical Emission Spectrometry (ICP- OES) |
AAS/ ICP-OES/ ICP- MS (wavelength 259.94) |
2. |
Milk |
Vitamin A |
AOAC 20th. Ed 2016 2001.13 |
Use of pyrogallic acid, saponify. Neutralize with acetic acid dilute with THF ethanol. Mobile phase: Methanol: water; isocratic) Gradient: Acetonitrile and Dichloromethane |
HPLC- UV |
|
|
Vitamin D2 |
AOAC 2016.05 |
|
Liquid Chromatography- Tandem Mass Spectrometry |
|
|
Vitamin D3 |
AOAC 2002.05 |
Sample size: 5-20g Ext. Solvent: Alcohol (Methanol/Isopropyl alcohol/Ethanol) Iso-octane etc. Mobile phase: Gradient (Water with Acetonitrile &Methanol) Gradient: Isopropanol-n-heptane (20+80). Dilute 200mL isopropanol to 1 L with n-heptane. |
HPLC- PDA/UV/ VWD |
3. |
Vegetable Oil |
Vitamin A |
AOAC 20th.Ed 2016 2001.13 |
Use of pyrogallic acid, saponify. Neutralize with acetic acid dilute with THF: ethanol. Mobile phase: Methanol: water; isocratic) Gradient: Acetonitrile and Dichloromethane |
HPCL- UV |
|
|
Vitamin D2 |
AOAC2016.05 |
|
Liquid Chromatography- Tandem Mass Spectrometry |
|
|
Vitamin D3 |
AOAC 2002.05 |
Sample size: 5-20g Ext. Solvent: Alcohol (methanol/Isopropyl alcohol/Ethanol) iso-octane etc. Mobile phase: Gradient (Water with Acetonitrile & methanol) Gradient: Isopropanol-n-heptane (20+80). Dilute 200mL isopropanol to 1 L with n-heptane. |
HPLC- PDA/ UV/VWD |
4. |
Atta |
Iron |
AOAC 2011.14 (ICP) AOAC 985.35 and AOAC 999.10 (AAS) AOAC 2015.06 (ICP-MS) |
Microwave assisted Inductively Coupled Plasma Optical Emission Spectrometry (ICP- OES) |
AAS/ ICP-OES/ ICP- MS (wave length 259.94) |
|
|
Vitamin B12 |
A2014.02, OAC 2011.10 & 2011.09- safety Precaution is to be taken while using KCN |
Sample extracted in KCN buffer (pH 4.5). Post autoclave and centrifugation, supernatant was syringe filtered prior to RP- HPLC using methanol -water gradient to resolve Vit.B12 peak. Sample concentration and purification using SPE further improved peak resolution and removal of interfering peaks. Use of Immuno-affinity cartridges help in selective concentration and extraction. Mobile phase: A) 0.025% TFA in water B) Acetonitrile |
HPLC/ UPLC UV (DAD) detector at 361nm
Use of Immunoaffinity column, NaCN followed by UPLC- UV
HPLC, requiring potassium cyanide |
|
|
Vitamin A |
AOAC 20th. Ed 2016 2001.13 |
Use of pyrogallic acid, saponify. Neutralize with acetic acid dilute with THF ethanol. Mobile phase: Methanol: water; Isocratic) Gradient: Acetonitrile and Dichloromethane |
HPLC -UV |
|
|
Thiamine |
AOAC 20th Ed. 2016, 957.17
|
Sample size; 5-10g Ext. process 0.1-0.5 NHCI/ NaoH/H2SO4/Phosphate buffer: Enzyme treatment: 0.1-1g takadiastase, papain, protease, etc. Mobile phase: Gradient (Phosphate buffer with Acetonitrile &Trifluoroacetic acid) |
HPLC-PDA/ FLD & LC-DAD/ MS (J.AOACInt 2009; 680-687) |
|
|
Riboflavin |
AOAC 20th Ed. 2016, 970.65 & J. Chro A, 2005, 49 |
Sample size; 5-10g Ext. process 0.1-0.5 NHCI/ NaoH/H2SO4/Phosphate buffer: Enzyme treatment: 0.1-1g takadiastase, papain, protease, etc. Mobile phase: Gradient (Phosphate buffer with Acetonitrile & Trifluoroacetic acid) |
HPLC-PDA/ FLD & LC-DAD/ MS (J.AOAC Int 2009; 680-687) |
5. |
Maida |
Same as Atta |
Same as Atta |
-do- |
-do- |
6. |
Rice |
Same as Atta |
Same as Atta |
-do- |
-do- |
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