Method Material Result

Enhanced  Extraction mediums were prepared using  Solvent mixture  yield of crude extracts
extraction yields mixtures of five components, (i) ethanol, (i) ethanol, (ii)  maximum yield values were obtained when extractions
and mobile (ii) ethyl acetate, (iii) dichloromethane, ethyl acetate, (iii) were performed with ethanol–dichloromethane (2.9288
phase separations (iv) acetone, and (v) chloroform, whose dichloromethane, g/20 g), ethanol–chloroform (2.9096 g/20 g) and ternary
by solvent proportions were varied according to a (iv) acetone, and ethanol–dichloromethane–chloroform (3.0247 g/20 g)
mixtures for simplex centroid design. (v) chloroform mixtures
the analysis of  Each extract was prepared by weighing  20g of dried and  yields of the neutral fraction
metabolites in 20 g of dried and crushed leaves and crushed leaves  Lower yield was observed with pure ethanol.
Annona adding 150 mL of the solvent mixtures,  ultrasonic bath  However, in the presence of ethyl acetate-
muricata L. leaves which then placed in an ultrasonic bath (Unique, model dichloromethane (1.9767 g/20 g), acetone–chloroform
2009 (Unique, model USC 1400) for 30 min USC 1400) (1.9006 g/20 g), ethanol–ethyl acetate–
aim of this work is  The bathwater was changed every 10 min  filter paper dichlorometane–chloroform (0.9356 g/20 g), and
to emphasize the to maintain constant water temperature.  weighed flask quinary
importance  The extracts were filtered through filter  HPLC analysis (1.9119/20 g70.0036) mixtures the yield increases
of the effects of paper to separate the solution from small  yields for organic fractions
solvents and their pieces of leaves and the solution was  ethanol (0.3894 g/20 g) resulted in effective increases
mixtures on the placed in an identified and weighed flask. in the extraction yields whereas the ethanol–
yields of metabolites  This procedure was repeated two more dichloromethane–acetone mixture showed the highest
in unfractionated times, so the total volume of solvent extraction efficiency (0.4259 g/20 g).
(crude) and mixture added to the leaves was 450 mL.  yields for basic fractions
fractionated  These solutions were left undisturbed  quaternary mixtures of ethanol– ethyl acetate–
extracts. under forced ventilation until reaching acetone–chloroform (0.4579 g/20 g), ethanol–ethyl
constant weight to remove residual acetate–dichloromethane–chloroform (0.6070 g/ 20 g),
solvent and quinary mixtures of the ethanol–ethyl acetate–
 Then, the 33 extracts were fractionated dichloromethane–acetone–chloroform mixture (0.5398/
by liquid–liquid extraction, resulting in 20 g±0.0539 g), increased the yield of this fraction
fibers and four more fractions  yields for the fiber fraction
denominated as neutral (fat and grease),  none resulted in high extraction yields for pure
organic (terpenoids and phenolic solvents.
compounds), basic (alkaloids) and polar  yield are obtained with ethyl acetate–dichloromethane
(quaternary alkaloids and n-oxide). The (0.5585 g), dichloromethane– chloroform (0.4171 g/20
residue is the fraction that remained g), ethyl acetate–dichloromethane– acetone–
after removal of the basic, organic, chloroform (0.4177 g/20 g), and ethyl acetate–acetone–
neutral, and fiber fractions. chloroform (0.4167 g/20 g) mixtures.
 yields for residual material
 highest yields are obtained with ethanol–dichloromethane
(0.7882 g/20 g), ethanol–chloroform (0.7526 g/ 20 g),
ethanol–dichloromethane–acetone (0.7117 g/20 g),and
ethanol–dichloromethane–chloroform (0.6749 g/20 g)
mixtures
Microwave  Extraction and fractionation  mechanical  Chloroform:Ethanol 3:2contained more amounts of
Assisted  leaves of Annona muricata were dried grinder the phenolic constituents, 360 mg GAE/100g
Extraction, under shade and then powdered with a  sieve No.42
Fractionation and  total flavanoid content of the extract is
mechanical grinder.  airtight
Total Phenolic and  The powder was passed through sieve container 97.5mgQE/100g
Flavanoid No.42 and stored in an airtight container  microwave oven
Estimation of for further use  ethanol
Annona muricata  The dried material was extracted in  Folin-ciocalteau
Leaves ethanol by scientific microwave oven reagent
2015  The extraction time was 5 minutes and  gallic acid
the power used was 850W  2% sodium
 About 30ml of ethanol was used as the carbonate
solvent  distilled water
 The extract was made solvent free by  sodium nitrite
distillation process and the resulting 5%
semisolid mass was vacuum dried to yield  10% aluminium
a solid residue.
chloride
 The experiment was repeated to get 20g
of the extract.  1M sodium
 Determination of total phenolic content hydroxide
 Folin-ciocalteau reagent and gallic acid
was used
 The reagent (5ml) was mixed with 1ml of
gallic acid at different concentrations and
after 3 minutes 4ml of 2% sodium
carbonate was added to each solution
 Determination of total flavonoids
 Total flavanoid content was determined
by aluminium chloride colorimetric assay
 Distilled water (4ml) was taken in 10ml
of volumetric flask and an aliquot
quantity of extract (1ml) or standard
solution of quercetin (25-150µg/ml) was
added to it
 Sodium nitrite 5% (0.3ml) was added
and after 5min,0.3ml of 10%
aluminium chloride was added.
 After 5 min, 2ml of 1M sodium
hydroxide was added and the volume
was made up to 10ml with distilled
water
UPLC–QTOF–  About 10 g of lyophilized leaves were mixed with
enough of HydroMatrixTM to fill the 100ml stainless
MS and NMR steel sample cell
 ethanol was added while the temperature and
analyses of pressure were increased to 100 ◦ C and 10.3 MPa
graviola (100 atm) over a 3 min time

(Annona
muricata)
leaves
2015