Detection of Adulteration in Ghee

Tanmay Hazra, Rohit G. Sindhavand D.C.Sen

INTRODUCTION

In India, Ghee is the second largest dairy product prepared from milk. Since the Vedic era it has an integral relationship with Indian culinary culture (Hazra, 2016). However, various ghee like products are very popular in different parts of world like Egypt (Samna), Sudan (samin), Uganda (samuli), etc. Moreover, it is gaining popularity in countries like Australia, United Kingdom, New Zealand, etc. (Parmar et al., 2018). Ghee is one of the superior edible fats that contains a plenty of bioactive components like conjugated linoleic acid (CLA), butyric acid, fat soluble vitamins (A and D), β-carotenes, etc. (Hazra, 2016). It was estimated that for 2020, combined butter and ghee (clarified butter) production was calculated to 6.1 million metric tons against 5.8 million metric tons in 2019 (Sharma et al., 2021). The applications of ghee is not restricted in food only. It has therapeutic as well as health benefit applications too (Hazra and Parmar, 2014).

Table 1: Chemical Composition of Ghee

Constituents	Cow milk ghee	Buffalo milk ghee
Fat (%)	99 - 99.5	99 - 99.5
Moisture (%)	<0.5	<0.5
Carotene (mg/g)	3.2 - 7.4	-
Vitamin A (IU/g)	19-34	17-38
Cholesterol (mg/100g)	302 – 362	209 – 312
Tocopherol (mg/g)	26 – 48	18 – 31
Free fatty acid (%)	2.8	2.8

ADULTERATION IN GHEE

Adulteration means “Fraudulent, intentional substitution or addition of a substance in a product for the purpose of increasing the apparent value of the product or reducing the cost of its production, i.e., for economic gain” (Hazra, 2016). Ghee is the costliest edible fat that is almost three to five times higher than commonly available different edible oils (cotton, coconut, palm or sunflower, etc.). The market demand of ghee is increasing day by day in national and international market. The market demands reaches highest especially during festive season in India. In this complex situation, some unscrupulous traders use to adulterate ghee with cheaper vegetable oils or fats. Reports have been appearing in the print and electronic media, indicating the rampant malpractices of ghee adulteration particularly, in the central and northern parts of the country. It is not known as to what extent these types of malpractices of adulteration are prevailing in the ghee trade in our country and what quality of ghee is available to the consumer. Different studies suggest that not only cheaper vegetable oils but also animal body fats are used to adulterate with pure ghee. In recent years, the problem of adulteration has turned a very serious dimension. In order to ensure a genuine product to the consumer, the Government of India has prescribed the compositional standards for ghee, under FSSR (2011). However, such rules can be violated by different means like using designer oils and fats etc. So, food researchers are always devoted themselves in search of different noble techniques for detection of adulteration in ghee efficiently. In the below, various techniques including their merits and demerits for detection of adulteration in ghee have been briefly discussed.

Table 2: FSSAI Standards of Ghee

Parameter	Ghee
Moisture, maximum, %, (m/m)	0.5
Milk fat, minimum, %, (m/m)	99.5
Butyro-refractometer Reading at 40ºC	Areas other than cotton tract areas: 40 - 43.5 b.      Cotton tract areas: 41.5 - 45
Reichert Meissl Value, minimum	Areas other than cotton tract areas: 28 Cotton tract areas: 24
Polenske Value	1.0 – 2.0
FFA (% Oleic Acid) (maximum)	3.0
Baudouin Test	Negative

(FSSAI, 2011)

MILK FAT CONSTANT ANALYSIS APPROACHES

Butyro-Refractometer Reading (BR reading)

The general values for BR reading (at 40°C) of pure milk fat ranges from (40–43) and using this method 10- 20% vegetable oil can be detected in ghee (Hazra, 2017). One of the drawbacks of the method is that the BR reading changes with the animal feed particularly cotton seed oil/cake which

limits its application (Sharma et al., 2020).

 

Reichert-Meissl (RM) Value

In India, RM value (measures butyric (C_4:0) and caproic (C_6:0) acids) analysis is widely applied to ascertain the purity of milk fat or ghee. For pure milk fat this value ranges from 28-32 (except in cottonseed area where it varies from 21-24) although, the lower RM value indicates milk fat adulteration with other foreign fats or oils (Ramani et al., 2019). Pranoti (2012) reported that pure cow ghee adulterated with depot body fat @10% level and in pure buffalo ghee adulterated with @ 20% depot body could be detected by RM value analysis. However, very recently Pathania et al., (2020) highlighted that this value can be modified by using different oils known as RM modifier oil; therefore, this analysis alone cannot ascertain the purity of ghee.

Polenske Value

Polenske value denotes majorly caprylic (C_8:0) and capric (C_10:0) acid present in milk fat or ghee.  This method can be used for the detection of adulteration of milk fat/ghee with coconut oil in ghee (Kumar et al., 2005).

Iodine Value

Iodine value is “Number of grams of iodine absorbed by 100 g of fat under specified conditions”. This constant is generally measures unsaturated fatty acids (double bond) present in milk fat or ghee. The iodine value for milk fat ranges from 26 to 35, which is low in comparison to most of the other fats and oils (Hazra, 2016). This technique can be successfully applied to detect hydrogenated vegetable oil in milk fat/ghee (Sharma et al., 2020); this method is also successfully applied to detect sheep body fat in ghee (@ 30%) (Gandhi et al., 2015). However, lower level of adulteration (less than 10%) in ghee cannot be detected with this technique (Ramani, 2019).

Saponification Value

Saponification value which denotes the number of milligrams of KOH required to saponify one gram of fat. It is an indication of average molecular weight of fatty acids present in milk fat or ghee. This method is suitable to detect mineral oil in ghee (Sharma et al., 2020).

METHODS BASED ON PHYSICAL PROPERTIES

Opacity Test

Opacity time of the normal pure milk fat/ ghee and animal depot fat are around 14–15 min and 10–20 s, respectively. Using this test protocol 5% level of animal body fat could be detected in ghee (Hazra, 2016). However, reproducibility is the major issue for this test protocol (Sharma et al., 2020).

Bomer Value

Bomer value is defined as sum of the melting point of saturated triglycerides (isolated from diethyl ether method) and twice the difference between this melting point and that of the fatty acids obtained after the saponification of these triglycerides. Using this test protocol @20% level of vegetable oil or animal depot fat could be detected in ghee(Sharma and Singhal, 1996). However, this protocol is tedious and time consuming, to perform specially in field condition.

Complete Liquefaction Time

The time taken by solid milk fat to become completely liquefied at a given temperature is defined as complete liquefaction time (CLT) and this technique has been used for the detection of adulteration in ghee. Upadhyay (2014) reported to detect 10% level of goat body fat in ghee with the use of this test protocol.

Apparent Solidification Time(AST)

The time taken by the melted fat to get apparently solidified at a given temperature is referred as apparent solidification time (AST).Gandhi et al., (2018) suggested to detect mixture of palm oil and body fat @ 20% level by applying this test protocol.

Chromogenic Test

Different quick chromogenic tests have been tried to identify the specific adulterant in ghee. In Table 3, a few such quick chromogenic tests have been shown.

Table 3: Chromogenic Test for Detection of Adulteration in Ghee

Sr. No	Name of the test	Types of adulterants detected	Observation
1.	Baudouin test	Vanaspati in pure ghee	Pink colour for adulterated ghee, colourless for pure ghee
2.	Halphen test	Cottonseed oil in pure ghee	Crimson colour observed for ghee adulterated with cotton seed oil
3.	B.N. Ghosh test	Vegetable oil	Appearance of a green colour confirms the presence of vegetable oil in ghee
5.	Potassium Ferricyanide and Ferric Chloride based chromogenic test	Palm oil in ghee	Deep green colour confirms the presence of palm oil in ghee
6.	DPPH (2,2-Diphenyl-1-picrylhydrazyl)(α,α-diphenyl-β-picrylhydrazyl)based chromogenic test	Palm oil in ghee	Violet colour confirms pure ghee; however, yellow colour confirms ghee adulterated with palm oil.

(Ramani, 2019)

However, cross reactivity with other minor components in ghee, temperature of storage may affect the results of these protocols. Hence, more researches are required for validation of these said chromogenic methods for successfully apply in field level conditions. Recently, different modern instrument based approaches have been applied for the detection of adulteration in ghee.

Tests Based on Gas Liquid Chromatography (GLC)

Fatty Acids Analysis by Gas Liquid Chromatography

Milk fat contains mainly short as well as medium chain fatty acids, while vegetable oils are rich in long chain unsaturated fatty acids. Therefore, the fatty acid composition using GLC has been used to detect vegetable oil in ghee (Upadhay, 2014).Hazra et al., (2020) reported to detect palm oil in ghee @ 5% by fatty acids analysis with GLC. Rebechi et al., (2016) advocated that fatty acids analysis combined with regression analysis, was a very authentic tool to ascertain the purity of milk fat.

Triglycerides(TGs)Analysis by Gas Liquid Chromatography

Hazra (2017)reported that both cow and buffalo ghee contained sixteen major TGs having carbon numbers varying from 24 to 54 but TGs having carbon numbers (C₃₆,C₃₈,C₄₀,C₄₂, C₅₀ and C₅₂) were predominant representing about 60% of total TGs present in both cow and buffalo ghee. Same author found that vegetable oils or body fats contained longercarbon number triglycerides but absence of shorter chain triglycerides. Based on these triglycerides ISO/IDF derived S-limits (S2, S3, S4 and S5) for the detection of different vegetable oils and animal fats in cow milk fat. The ISO/IDF reference values for cow milk fat are ST: 95.68 – 104.32; S2(Soya bean, Sunflower, olive, Rape-seed, Linseed, Wheat germ, maize germ, Cotton seed & Fish oil): 98.05 – 101.95; S3 (Coconut & Palm kernel fat): 99.42 – 100.58; S4 (Palm oil & beef tallow): 95.90– 104.10; S5 (Lard) : 97.96 – 102.4). AmruthaKala (2013) developed a test to detect very low level of adulteration in ghee and few Indian dairies adopted this test protocol and standard to detect presence of adulterants in milk fats or ghee. However, Sharma et al., (2021) claimed that this standard was not suitable in pan India basis. Hence, this protocol is not suitable to detect adulteration of foreign fats in ghee.

Tests Based on High-Performance Liquid Chromatography (HPLC)

Raniet et al., (2015) advocated that using HPLC, the un-saponifiable matter of ghee analysis (β-sitosterol as marker),1% vegetable oil in ghee could be detected. This technique has been recently adopted by FSSAI to ascertain the purity of ghee.

Tests Based on Reverse Phase Thin Layer Chromatography (RP-TLC)

IDF (1965) recommended a RP-TLC based method for the detection of vegetable fats in milk fat on the basis of the appearance of a small band of β-sitosterol in addition to the major band of cholesterol (Hazra, 2016). Recently, Rani et al., (2013) and Upadhyay (2014) reported that using RP-TLC, even up to 1% vegetable oil could be detected in milk fat or ghee using β-sitosterol as biomarker.

Tests Based on Spectroscopic Methods

Jha (1981) applied visible spectroscopy for the detection of Cheuri (Madhuca butyracea) fat in ghee, a common adulterant in Nepal. Researchers developed methods using fourier transform - near infrared (FT-NIR) and fourier transform - middle infrared (FT-MIR) spectroscopy to identify palm oil adulteration in ghee (Aparnathi et al., 2019).

Tests Based on DNA Analysis

Hazra (2016) was able to isolate DNA from ghee and amplified those DNA by simple PCR and those researchers claimed to detect goat body fat adulteration in ghee. In a similar research Sharma et al., (2018) were able to detect 10% buffalo tallow in cow ghee.

CONCLUSION

Ghee, is one of the superior edible fats that contains various bioactive components; so, apart from culinary use it has plenty of health benefits too. However, this image of ghee is being tarnished due to the evil practices like adulteration with inferior qualities of fats and oils with ghee. Different analytical techniques have been applied for detection of adulteration in ghee. However, no single analytical method can ascertain the purity of ghee totally. Therefore, FSSAI has recommended a combination of different test protocols to detect the adulteration of ghee. Side by side, it is suggested that both government and food manufacturers (small or big) jointly should come forward to take the necessary steps for solving this problem of adulteration. Further, government should also promote the growth of organized dairy business that will be beneficial for both consumer and Indian dairy industry in near future.

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