An Investigation into Phytochemical Profile and Nutraceutical Value of Amla (Emblica officinalis) Fruits

Abstract

Recently, nutraceuticals and medicinal plants have become the focus of current medical research. The fruit or fruit pulp of Emblica officinalis is a reputed drug of Ayurvedic, Unani, Siddha and Homoeopathic systems of medicine and believed to increase defense against diseases. The fruit primarily contained tannins, alkaloids, phenolic compounds, amino acids, carbohydrates and other compounds especially the essential nutrients. The vitamin C content in Amla varies from 200-900mg/100g from batch to batch. The comprised investigation explored physico-chemical, phyto-chemical and nutraceutical profile of this versatile and reputed ethnomedicinal fruit through basic scientific research to establish the rational scientific foundation of its utility as rejuvenating food and tonic.

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Int J Mod Pharm Res/Vol 2/Issue 1/Jan-Feb 13
Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
Volume 1, Issue 4
AN INVESTIGATION INTO PHYTOCHEMICAL PROFILE AND
NEUTRACEUTICAL VALUE OF AMLA (EMBLICA OFFICINALIS)
FRUITS
Nikhil K Sachan
1
,
Sudhir Singh Gangwar
2
, Ranjana Sharma
2
, Yatindra Kumar
2*
1
University Institute of Pharmacy, C.S.J.M. University, Kanpur – 208 024 Uttar Pradesh
2
Dept. of Pharmacy, G.S.V.M. Govt. Medical College, Kanpur – 208 002 Uttar Pradesh
Received 12 Feb 2013; Revised 20 Feb 2013; Accepted 25 Feb 2013
Key words: Indian gooseberry, food supplement, medicinal foods, emblic myrobalan,
healthcare
INTRODUCTION
Fruits are amongst the first food items known to human beings. Fruits, whether fresh or dried,
have always formed a part of the staple diet of human beings
[1]
. The reason for this is that
they are rich in nutrients and provide some of the essential minerals, vitamins, and the like, to
our body. Apart from that, they also help in curing a number of diseases. Amla consist of
fress or dried fruits of Emblica officinalis; it is one of the important herbal drugs used
traditionally both as a medicine and as a tonic to build up lost vitality and vigor
[2]
. In Unani
medicine, it is described as a tonic for heart and brain. According to the two main classic
texts on Ayurved, Charak Samhita and Sushrut Samhita, Amalaki is regarded as “the best
among rejuvenative herbs”, “useful in relieving cough and skin disease” and “the best among
the sour fruits”
[3]
. Amla grows in tropical and subtropical parts of India, China, Indonesia,
INTERNATIONAL JOURNAL OF
MODERN PHARMACEUTICAL RESEARCH
Research Article
ABSTRACT
Recently, nutraceuticals and medicinal plants have become the focus of current medical research. The
fruit or fruit pulp of Emblica officinalis is a reputed drug of Ayurvedic, Unani, Siddha and
Homoeopathic systems of medicine and believed to increase defense against diseases. The fruit
primarily contained tannins, alkaloids, phenolic compounds, amino acids, carbohydrates and other
compounds especially the essential nutrients. The vitamin C content in Amla varies from 200-900mg
/100g from batch to batch. The comprised investigation explored physico-chemical, phyto-chemical
and nutraceutical profile of this versatile and reputed ethnomedicinal fruit through basic scientific
research to establish the rational scientific foundation of its utility as rejuvenating food and tonic.
ISSN
2319 – 5878

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
and the Malay Peninsula
[4, 5]
. The fruit is spherical pale yellow with six vertical furrows. The
average weight of the fruit is 40-50g. The fruits of Amla are widely used in the Ayurvedic
preparations and are believed to increase defense against diseases. Fresh fruit is refrigerant,
diuretic and laxative. Fruit is also carminative and stomachic. Dried fruit is sour and
astringent. It is an important dietary source of vitamin C, amino acids, and minerals. Amla is
presently underutilized fruit, but has enormous potential in the world market. It is almost
entirely unknown in the world market and needs to be popularized. The export of amla fruits
from India, reported to be about 500 tons. It is being exported under the category of
Ayurvedic and Unani herbs
[6]
. Its medicinal and nutritional properties and culinary uses need
to be highlighted.
A nutrient is defined as a chemical whose absence from diet for a long enough time results in
a specific change in health. We need nutrients for normal body growth and development, for
maintaining cells and tissues, for fuel to do physical and metabolic processes. Foods provide
six major classes of nutrients viz., carbohydrates, lipids, proteins, minerals, vitamins and
water. The first five are called essential nutrients. Carbohydrates, protein and fats are called
macronutrients because our bodies need large quantities of them. The science of nutrition
helps us to improve our food choices by identifying the amounts of nutrients we need, the
best food sources of those nutrients we need, and other components in foods that may be
helpful or harmful. Learning about nutrition will help us make better food choices, improve
about their health and look, the market of dietary supplements and sport nutrition products is
expected to grow faster. The neutraceutical market has been growing at compound annual
growth rate (CAGR) of 20 percent for the last three years. The Indian nutraceutical market is
estimated to grow to $2731 million to 2016 at a CAGR of 13 percent. The Indian
pharmaceutical market is dominated by the pharmaceuticals and Fast Moving Consumer
Goods (FMCG) giants. The Indian nutraceutical market was $1180 in the year 2011
[7]
. The
our health, reduce the risk of many chronic diseases and increase our longevity. The term
nutraceutical is a portmanteau word coined with “nutrition” and “pharmaceutical” which
refers to a foodstuff (as a fortified food or dietary supplement) that provides health benefits.
The American Neutraceutical Association defines it as ‘a nutraceutical, is any substance that
is a food or a part of a food and provides medical or health benefits, including the prevention
and treatment of disease. Such products may range from isolated nutrients, dietary
supplements and specific diets to genetically engineered designer foods, herbal products, and
processed foods such as cereals, soups and beverages’. As Indians are becoming conscious

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
dietary supplements constitutes largest category accounting for 64% of nutraceutical market
driven primarily by pharmaceutical sector in the form of vitamins and mineral supplements.
It is thus becoming a logical progression for many food companies to enter into nutraceutical
market. The trend towards preventive healthcare have been observed with many
pharmaceutical companies including Novartis, GlaxoSmithKline and Cadila Healthcare etc
which have diversified into the production of nutraceuticals. Then there are fast moving
companies like Cadbury India and Dabur that have had the presence in the market with a slew
of supplements and additives. On the end of the spectrum companies that specialize in
ayurvedic / herbal healthcare products like the Himalaya Drug Company
[8]
. The Big global
players in which are always on lookout for ways to diversify their products line, have set up
the functional food or nutraceutical divisions; the pharmaceutical companies are also now
adopting the nutraceuticals and recent trends is convergence of food manufacturing
companies with pharmaceuticals to implement the research strategy. The evidence based
investigation concept called positivism play a greater role in the pharmaceutical and
nutraceutical research. In fact the humans, throughout the history, have been with noticeable
concern in the healthcare, though the concepts themselves took a very long time to develop
into a body of knowledge. A logical approach to the study of drugs and their activities is the
recognition of the basic principles behind the biochemical events leading to drug actions.
Nowadays, an increasing amount of insight into the behavior of drugs at the macromolecular
level has been developed and there is a lot of direct and indirect evidence supporting these
biochemical postulations of drug action. However, in the so-called pre-scientific period,
natural products have a history of therapy in the form of folk remedies, but little of today’s
drug therapy is based on these remedies. Some of the natural products currently used, either
as such or as derivatives may often be used originally for other purposes, such as arrow
poisons, part of religious or other rituals, and even cosmetics. An intensive and extensive
study of the naturally occurring molecules identified as ‘therapeutically active’ is desired
urgently to come out with new therapeutic entities. Similarly, recognising the merits of a
wider usage of certain fruits and botanicals have triggered their scientific validation towards
intended nutraceutical utility. Therefore these products, after having proven their ability of
being fortified food supplements towards preventive healthcare, under the light of modern
scientific principles, can be formulated to regulated markets, and simultaneously can boost
the sale in domestic market convincing the educated consumers who are the major buyers of
these products.

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
MATERIALS AND METHODS
Collection and Processing of the Materials
The fresh fruits of Amla were purchased from open market at district Kanpur of Uttar
Pradesh state. The fruits were processed to remove the fruit pulp for preliminary physico-
chemical analysis and isolated pulp was homogenized and passed through a strainer for
further subject to spray drying obtaining dry powder. Fresh fruit pulp was analysed for pH,
moisture content, titratable acidity, ash and total water-insoluble solids according to
Association of Official Analytical Chemists (A.O.A.C.)
[9]
. For the spray drying, the
homogenized juice was evaporated to about 50% with a rotary vacuum evaporator and mixed
with maltodextrin (5% w/v of initial juice) followed by spray drying using spray dryer at
200°C inlet temperature and 150°C outlet temperature with optimized aspiration speed to dry
the sample. The powder was stored in an airtight container and kept in desiccators till further
use.
Moisture Content Determination
The moisture content of the sample was estimated using the Karl-Fischer titration apparatus
using standard KFR reagent.
Minerals and Crude Fat Estimation
The determination of minerals and crude content of the samples were determined as per
AOAC (1997) procedures; whereas minerals (P and Fe) were estimated as per the AOAC
(1990) procedures. Calcium, Sodium and Magnesium were estimated using the flame
photometry.
Total Phenolic Content
TPC was determined using the Folin-Ciocalteu’s reagent using the method as per Lako et al.
(2008)
[10]
. Samples of 0.3 ml were introduced into test tubes followed by 1.5 ml of Folin-
Ciocalteu’s reagent (diluted 10 times with water) and 1.2 ml of sodium carbonate (7.5% w/v).
The tubes were vortexed, covered with parafilm and allowed to stand for 30 min. Absorption
at 765 nm was measured. If the sample absorbance exceeded 1, the sample was appropriately
diluted to give a reading of less than 1. Total phenol contents were expressed in gallic acid
equivalents (mg per 100 gram fresh fruit). The gallic acid standard line has the equation y =
0.0111x - 0.0148 (R2 = 0.9998), where y is absorbance at 765 nm and x is concentration of
gallic acid in mg/L. All Absorptions were measured in three replicates.

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Detection of Carbohydrates
Extracts were dissolved separately and were tested with Molisch reagent, Fehling’s reagent,
Bendict solution, and Barfoed’s test for detection of carbohydrates
[11]
.
a) Molisch’s test: To the filtrate, added few drops of alcoholic alpha nepthol and 2ml of
concentrated sulfuric acid slowly through the side of test tube; presence of
carbohydrate produce a violet colour ring at the junction of two layers.
b) Fehling’s test: A little fraction of filtrate treated with Fehlin’s solution I & II and then
heated on a water bath. A brick red precipitate is indicator for reducing sugars.
c) Bendict’s test: Small quantity of filtrate treated with equal quantities of Bendict’s
reagent, heated subsequently on a water bath result to formation of a brown
precipitate in presence of reducing sugars.
d) Barfoed’s test: The different extracts were treated with Barfoed’s reagent.
Monosaccharides, if present, produce a brick red precipitate.
Detection of Glycosides
Glycosides were confirmed by subjecting the acid hydrolysed extract to Legal’s test,
Borntrager test and Libermann-Burchard’s test
[11]
.
a) Legal’s test: Hydrolysate was dissolved in pyridine and sodium nitro-prusside
solution, added sodium hydroxide; a colour change result in presence of glycosides.
b) Borntrager’s test: A few milliliters of hydrolysate treated with chloroform, decanted
off chloroform layer, added equal quantity of dilute ammonium solution. A pink
colour is produced in ammonical layer in presence of glycosides.
c) Libermann-Burchard’s test: Hydrolysate treated with chloroform, to this added
Libermann-burchard reagent; a colour change result in presence of glycosides.
Detection of Fixed oil and Fats
[11, 12]
a) Spot Test: A small quantity of sampler was separately pressed between two filter
papers and was observed for appearance of oil stain on paper which will indicate
presence of fixed oils or fats.
b) Saponification test: Few drops of 0.5N potassium hydroxixe along with one or two
drops of phenolphthalein were added to various extracts, heated on a water bath for 1-
2 hours. Saponification or no saponification indicates the presence or absence of oil
and fats.

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
Detection of Protein & Amino acids
The spay dried extract was subject to Million’s test reagent, Biuret test reagent, and Nin-
hydrin test reagent, presence of amino acids and proteins is indicated by production of red,
violet and blue colour respectively
[11]
.
Detection of Phenolic compounds & Tannins
All the dry extracts were dissolved in minimum amount of water, filtered and subject to
Ferric chloride test, Gelatin test. Filtrate on addition of few drips of ferric chloride produce a
violet colour precipitate in presence of tannins. A white precipitate is resulted in presence of
tannins on addition of 1ml 1% solution of gelatin to the filtrate
[11-13]
.
Table 1: Preliminary Macroscopic and Physical Examination of E. officinalis
Fruit Shape
Globular
Size of Fruits
3.8cm (Average)
Fruit Weight
41.24 – 52.48g
Colour of Fruits
Greenish Yellow
Sp. Features
Segmented through 6 ridges
Pulp Weight
91% of Fruit Weight
pH
3.1 – 3.4
Seeds
Six
Odour
Astringent & Sour
Taste
Characteristic
(sweet, sour, pungent,
bitter and astringent)
Spray Dried Powder
Off-white Cream Colour to
Brownish Yellow
Water Soluble Solids
≥28 % w/w
Foreign Matter
< 1 % w/w
Moisture Content
8.1 % w/w
Ash Value
< 8 % w/w
Acid Insoluble Ash
< 1.1 % w/w
Organic Extractives
28 – 30 % w/w
Sand / Silica
Absent

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
Detection of Phytosterols
Small quantity of the dry extracts dissolved in about 5ml of the chloroform and subjected to
Salkowski’s test and Libermann-Burchard’s test
[11-13]
.
a) Salkowski’s test: one ml of the chloroform solution, prepared as above was added
with few drops of concentrated sulfuric acid; green colour is the indicative of
phytosterols.
b) Libermann-Burchard’s test: The chloroform solution, prepared as above was treated
with few drops of concentrated sulfuric acid followed by one ml of acetic anhydride.
Presence of phytosterols is confirmed by the production of a bluish green colour.
Detection of Alkaloids
Small fractions of solvent free extracts were separately stirred with a milliliters of dilute
hydrochloric acid and filtered, the filtrate is tested with Mayer’s reagent, Wagner’s reagent,
Hanger’s reagent, Dragendroffs reagent to confirm the presence or absence of alkaloids as
indicated by production of cream, reddish brown, yellow or brown colour respectively with
these reagents in presence of alkaloidal substances
[11, 13]
.
Detection of Flavonoids
Shinoda’s test – Small quantity of the extract was dissolved in alcohol, to those pieces of
magnesium followed by concentrated hydrochloric acid was added drop wise and heated.
Appearance of magenta color shows the presence of flavonoids
[12,13]
.
Table: 2: Composition of fruit pulp of Emblica officinalis
Composition
Percentage
Moisture
81.1%
Protein
0.5%
Fat
0.1 %
Mineral Matter
0.7%
Carbohydrate
14.2%
Fibre
3.4%
Calcium
0.05
Phosphorus
0.02
Iron
1.2 mg/100g
Nicotinic Acid
0.2 mg/100g
Vitamin C
600mg/100g

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Detection of Saponins
Foam test – The extract was diluted with 20 ml of distilled water and it was agitated in a
graduated cylinder for 15 minutes. Formation of 1 cm layer of foam shows the presence of
saponins
[12]
.
Detection of Gums and Mucilage
Small quantity of the extracts were added separately to 25 ml of absolute alcohol with
constant stirring and filtered. The precipitate was dried in air and examined for its swelling
properties. No swelling was observed indicates the absence of gums and mucilages.
Precipitate tested for the swelling and presence and carbohydrate
[11]
.
Estimation of Ascorbic Acid
The determination of ascorbic acid content was performed using the method described in
Indian Pharmacopoeia. Sample solution equivalent to 0.2 mg ascorbic acid/ml was prepared
in water containing 3% (w/v) metaphosphoric acid. It was titrated against standard 2, 6
dichlorophenol 8ndophenols (2,6 DCIP) solution of 0.5 mg/ml concentration until the pink
color developed completely. The operation was repeated with a blank (IP, 1966)
[14]
.
RESULTS AND DISCUSSION
The fresh fruits of Amla were of about three centimeter average length and about 3.8 cm in
diameter. All the fruits were smooth segmented into six almost equal segments with formed
ridges extending from apex to base; the fruits were ranging from 21.24 to 32.48 gram in
weight (Fig. 1). The preliminary macroscopic properties and observations of physical
examination of fresh fruits and spray dried juice/extract are tabulated in Table 1. The fruits of
Emblica officinalis are rich in tannins. The fruits have 28% of the total tannins distributed in
the whole plant. The Amla is reported to contain two hydrolysable tannins Emblicanin A and
B, which have antioxidant properties, one on hydrolysis gives gallic acid, ellagic acid and
glucose wherein the other gives ellagic acid and glucose. Emblica fruit is a rich source of
vitamin C (ascorbic acid); a small fruit contains more vitamin C than a large fruit.
The fruit also contains considerably higher concentration of most minerals calcium,
phosphorus, iron and nicotinic acid along with amino acids (Table 2). The tests for
aminoacids have been identified positive; it is reported to have variety of amino acids viz.
glutamic acid, proline, aspartic acid, alanine, and lysine etc. Qualitative chemical
examination of the dried fruit powder of Emblica officinalis revealed the presence or absence

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
of various plant constituents. The observations were recorded in + (present) or – (absent) and
are summarized in the Table 3.
Table: 3: Phytochemical Screening of Emblica officinalis
Constituents
Tests/Reagents
Observations
Alkaloids
o Hager’s Reagent
+
o Wagner’s Reagent
+
o Mayer’s Test
+
o Dragendorff Test
+
Carbohydrate
o Molisch’s Reagent
+
o Fehling Solution
+
o Benedict’s Reagent
+
o Iodine Test
+
o Borntrager
+
Glycosides
o Legal’s Test
+
o Liberman Burchard's Test
+
Fixed Oils and Fats
o Saponification Test
−
o Spot Test
−
Proteins and Amino Acids
o Millon’s Reagent
−
o Biuret Test
+
o Nin-hydrin Test
+
Phenolic Compounds
o Neutral FeCl
3
+
o Gelatin Test
+
Saponins
o Foam Test
−
Phytosterols
o Liberman Test
−
o Salkowski test
+
Gums and Mucilages
+
Amla is one of the most celebrated herbs in the Indian traditional medicine system, and is
becoming increasingly well known for its unusually high levels of Vitamin C, which is
resistant to storage and heat damage due to cooking. This fruit is native to India as it is self

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
evident from the English name ‘Indian Gooseberry’. It has found its use in various herbal
products for medicinal and nutritional value.
Eating the fruits and vegetables" is one of the tried and true recommendations for a healthy
diet. The edible portion of most types of fruits contains 75-95% of water. Fruits usually are
low in calories and with the exception of avocados and olives, contain practically contain no
fat. More than 900 different phytochemicals have been identified as components of food, and
many more phytochemicals continue to be discovered today. It is estimated that there may be
more than 100 different phytochemicals in just one serving of vegetables/fruit bite. These
popularly known as antioxidants can scavenge or mop off the harmful free radicals produced
in the body. Free radicals
damage cellular membranes,
proteins and DNA and cells
and produce a range of
diseases in body which could
be prevented with such fruits
consumption. Phenols,
flavonoids, anthocyanins and
carotenoids are some of the
important antioxidant found in
fruits and vegetables. The
study has explored nutrient
and non-nutrient components of E. ofiicinalis. As the results revealed that the Amla has
important nutritional factors contained with minerals, amino acids and vitamin. Its proven
nutraceutical potential will serve the popularity of fruit in the world wide educated
population. The vitamin C in Amla is remarkably stableeven after prolonged cold storage or
cooking, probably due to the presence of phenolic compounds tannins and phytophenols
which retarded the oxidation of ascorbic acid present in the fruit. Amla has been and remains
a product of substantial interest in India and south asia. It is particularly noteworthy that amla
is reported as an essential component in rasayanas such as triphala and chyawanprash to assist the
body attain a state of improved nutrition that prevents aging and enhances longevity.
Indian
Herbals Research and Supply Co. Ltd, furnished information submitted by GRAS Associated
to USFDA on behalf of Natreon Inc for consideration of Amla products under ‘Generally
Regarded as Safe’ Category and exemption from premarketing approval for its products
[15]
.
Figure 1: Fresh Fruits of Emblica officinalis

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Kumar Y et al.; Phytochemical profile and neutraceutical value of amla fruits
Many of investigations and traditional literature has been cited for the purpose. In fact this
will help the Amla fruit to get potential market abroad in addition to India. So the present
investigation will lay supportive base for the nutraceutical utility of Indian Gooseberry in
international market.
CONCLUSIONS
Amla, is an important medicinal plant of India. Its fruits have potent antioxidant activity due
to the presence of tannoids, tannins, vitamin C and flavonoids. It has been used as
rejuvenating herb. The observations in present investigation indicated that the emblic fruit is
highly nutritious. With the ease of long distance transportation with high degree of stable
Vitamin C in storage and thermal treatments, it servs as the packets of vitamin C with
wrapped with essential minerals amino acids and other vital nutrients.
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