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Procedia Chemistry 13 (2014) 57 62

International Seminar on Natural Product Medicines, ISNPM 2012

Changes in Secondary Metabolite Contents Following Crude


Drug Preparation
Sukrasnoa*
a

Pharmaceutical Biology Research Group School of Pharmacy, Insttut Teknologi Bandung


Jl. Ganesa No. 10 Bandung, Indonesia

Abstract
Crude drug is commonly prepared by directly drying of freshly harvested plant organ or after slicing. These processes may
decrease or maintain the content of the desired metabolites presence in the crude drug. Plant rhizome can be directly sliced
followed by drying or after storage at certain period. Certain rhizome can maintain the secondary metabolites after being stored
for three months (12 weeks), while others decreased just after being stored for two weeks. Drying process can be performed
under the sun or in an air circulated oven with temperature not higher than 60 qC. Phenolic content of crude drugs on the other
hand is the lowest if it is dried at 60 qC. Drying at 40 qC, 80 qC and 100 qC produce crude drug with higher phenolic content
compared to those dried at 60 qC. This may associated with the activity of peroxidase that has optimal activity at 60q C. At above
60 qC, the activity of peroxidase may decrease due to the degradation of the enzyme. Moist treatment of fresh material may
increase the content of the secondary metabolites. Boiling of Cosmos caudatus leaves increased the content of the flavonoid
glycoside. However, part of the flavonoid was presence in the aliquot that hamper further step of crude drug preparation.
Steaming of potato peels increased the chlorogenic acid content. From these observations, steaming can be considered as one of
pre-treatment steps in the preparation of crude drugs prior drying process. The increase of flavonoid glycoside in Cosmos
caudatus leaves upon boiling has been confirmed not due to the increase the extractability of the flavonoid. The increase of key
enzyme activity that involved in the biosynthetic pathway upon moist-heat treatment need to be further studied

Published
by Elsevier
B.V. B.V.
This is an open access article under the CC BY-NC-ND license
2014
2014The
TheAuthors.
Authors.
Published
by Elsevier
(http://creativecommons.org/licenses/by-nc-nd/3.0/).
Peer-review under responsibility of the School of Pharmacy, Bandung Institute of Technology.
Peer-review under responsibility of the School of Pharmacy, Bandung Institute of Technology
Keywords: representative secondary metabolite, storage, phenolic content, flavonoid glycoside, chlorogenic acid, boiling, crude drug processing

* Corresponding author. Tel.: +62-22-2508143; fax: +62-22-2508143.


E-mail address:sukras@fa.itb.ac.id

1876-6196 2014 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license
(http://creativecommons.org/licenses/by-nc-nd/3.0/).
Peer-review under responsibility of the School of Pharmacy, Bandung Institute of Technology
doi:10.1016/j.proche.2014.12.006

58

Sukrasno / Procedia Chemistry 13 (2014) 57 62

1. Introduction
Crude drug is commonly prepared in dry form in order to assure the stability, standardization and effective
handling such as transportation and storage. When plant organ is harvested from the intact plant, biochemical
reaction is still actively taking place. However the reaction is also strongly influenced by the environment such as
temperature, humidity and microorganism. Some bulky fresh plant organs can be stored for certain period without
significant change in quality. Improvement of quality may take place during storage due to fermentation process and
initiate the formation of the desired metabolites such as the release of aglycones, the development of flavor, etc.
However, many of them need to be further processed as soon as possible to produce high yield of the end product.
Pre-treatments before slicing and or drying may be introduced in the preparation of crude drugs. Such treatments
can increase the content of the desired metabolites, however, in other cases decrease or even eliminate certain
metabolites accompanied by the formation of new metabolites. Heat treatment is introduced in the preparation of
food products. Fermentation and drying at high temperature changed polyphenolic content and improve the flavour
of tea leaves. Following a series treatment from one kind of tea leaves can produce 16 different classes of tea.
Roasting of coffee bean eliminate chlorogenic acid that present in high quantity accompanied by the change in color,
the increase of bean fragility and the appearance of distinct coffee flavor. The chlorogenic acid in coffee beans was
converted into chlorogenic acid lactones during roasting1. In this paper, the influence of some treatments in crude
drug preparation such as storage of bulky organ, heat treatment, slicing and grinding, on the quality of crude drug is
reviewed.
2. Storage of Bulky Organ
Spices derived from rhizomes such as turmeric, ginger, galanga, campheria, etc. are frequently commercialized in
fresh and intact form. Since the metabolisms are still active, storage of bulky organs will definitely alter the patern of
metabolites present in the organ and influence its quality. Sugar cane stems need to be processed after harvesting,
otherwise the sucrose will be metabolized and sugar yield substantially decrease. Tobacco leaves need to be directly
sliced and dried after harvesting, delay in processing will deteriorate the quality of the tobacco produced. Similar to
tobacco leaves, the tuber of Amorphophalus oncophilus need to be sliced and dried after harvesting, delay in
processing will make the material become tough, viscid like concrete and create difficulty in grinding.
Storage of tobacco leaves at high humidity increase nitrite, alkaloid and nitrosamine specific tobacco within 7
days and these compounds decrease dramatically after 21 days. While storage at lower humidity decrease the
alkaloid but no increase in nitrite or tobacco nitrosamine and the amount remain constant after 28 days2. The
decrease of epicatechin and polyphenols in cocoa during fermentation is due to the diffusion out from the
cotyledons, polyphenol oxidation, and condensation. The flavors of cocoa is initiated during fermentation and further
developed when beans are dried 3. Storage of potato tubers decrease the carotenoid contents, especially carotenoid
derived from E-carotene and the decrease is higher at lower temperature. Exposure to mercury or sodium light
increase total carotenoid contents4. These observations suggest that manipulation of storage condition may enhance
the desired metabolites present in the crude drugs.
Ginger (Zingiber officinalis) can be stored for one week without significant change in the quality and the quantity
of volatile oil. Storage of ginger rhizome for one week will significantly reduce the moist present in the rhizome and
increase the yield of volatile oil during steam distillation compared to fresh rhizome. However, after two weeks,
shoots will grow from the rhizome and the oil content drastically decrease. Not only the content of oil, change also
takes place with the composition of the components present in the volatile oil5. Therefore, ginger oil taster can
distinguish the quality of ginger oil base on the period of bulky ginger storage. Similar pattern can be observed with
the oil content of Curcuma upon storage of bulky rhizome. However, Curcuma rhizome maintains the curcuminoid
content at approximately 2 % after being stored for 12 weeks 6. The volatile oil content however decrease
significantly from 1.10 % in fresh rhizome to 0.47 % after being stored for three months7. It is very likely to obtain
maximal yield of volatile oil from the rhizome of Zingiberaceae, the oil may be best obtained from fresh rhizome or
those that have been stored for not longer than two weeks. Unfortunately, spices from Zingiberaceae are frequently

Sukrasno / Procedia Chemistry 13 (2014) 57 62

59

used in fresh rhizome but may have been stored as the whole rhizome for long period.
3. Heat Treatment
Many vegetables contain flavonoid that has antioxidant activity. Various cooking methods may be involved
before being served, such as: boiling, sauting, steaming, frying, air oven heating or without any heat treatment.
Experiments on the effect of boiling on the content of flavonoid contents have been conducted toward 17 vegetables.
The decrease of flavonoid content was observed in two vegetables, remain constant in eight vegetables and increase
in seven vegetables. In general boiling does not decrease the flavonoid content. In certain species such as Cosmos
caudatus leaves and water spinach leaves, substantial increase of flavonoid content takes place upon boiling.
Boiling has been reported to substantially increas flavonoid content of Cosmos leaves 8. Kaempferol 3-Ogentiobioside substantially increase in heat treated Cassia alata leaves. Boiling increases the carotenoid of broccoli
and the polyphenols of fresh Brussel sprout9. The total antioxidant capacity of extract from various vegetables also
increase after boiling and steaming10. Heat treatment was conducted by treating in hot water at 85 qC for 4 minutes
followed by cutting and further treatment at 40 qC under high humidity for 4 hours followed by drying under the
sun. Direct sun drying on the other hand decrease and even remove glycoside content within 60 minutes 11. Such
increase may be due to the increase of flavonoid extractability or de novo formation during boiling process.
Experiment by boiling of Cosmos caudatus leaves in water and in 70% methanol was performed to test the above
possibility. In this experiment, direct extraction was performed by grinding fresh tissue with 70% MeOH followed
by maceration for 24 hours and rinsing the marc three times with the same solvent in order to exhaustively extract
the whole flavonoids present in the leaves. This extraction procedure is matched with those recommended by Xu et
al. (2005)12. Extraction through boiling was conducted by boiling 5 g intact leaves in 50 mL solvent for 15 minutes
followed by analysis of flavonoids from the aliquot and the marc. Flavonoid from the marc was analyzed in the same
way as that of extraction from fresh tissue. It was found that that boiling in MeOH 70% yielded 11.91 mg/100 g of
total flavonoid while boiling in water yielded 26.23 mg/100 g or increase by 120.23% 8. Therefore, it is very likely
that such increase is not due to the enhancement of flavonoid extractability but de novo formation of flavonoid
during the heating of the tissue in the aqueous environment. In young tissue, common precursor for the synthesis of
flavonoid together with other phenylpropanoid are available and upon heating the activity of enzymes involved in
flavonoid synthesis may increase and rapid conversion of precursor to flavonoid taken place. In addition, boiling
temperature will inactivate polyphenoloxidase13. Further study is needed to provide further support and to study the
mechanism involved. High temperature treatments will mostly inactivate the native enzymes; however, it will also
affect the texture of the material and the stability of the metabolites, especially thermolabile compounds.
Heating treatment influences phenolic content in other plant materials. It has been demonstrated that roasting
increase total phenol and procyanidin in peel almond14. Other researches demonstrated the decrease of total penolic
content and the antioxidant capacity of Alpinia zerumbet, Etlingera elatior, Curcuma longa and Kaempferia galanga
leaves upon drying on air, microwave oven and air oven at 50 oC. Freezedrying produced higher total phenolic
content compared to fresh. Freeze-dried product can be stored without significant decrease in total phenolic content
and antioxidant activity15.
Murakami et al. studied the influence of heating on the stability of flavonoids. Flavonoid such as rutin, luteolin,
luteoin-7-glucoside and chlorogenic acid gradually decomposed when heated at 100 qC16. In the presence of
chlorogenic acid the decomposition of rutin was inhibited at 100 qC. Rapid degradation takes place at 180 qC but
the decomposition product still has radical scavenging activity. Purple willow (Salix purpurea) freeze-drying of leaf
with first frozen with liquid nitrogen produce substantial change in flavonoid and salycilates qualitative and
quantitatively. Freeze drying without pre-freezing and vaccuum drying produce similar phenolic compounds as that
of fresh. Heat drying 60 qC and 90 qC induced substantial change in phenolics and mostly decrease except apigenin.
Most o-dihydroxy phenolic decrease upon drying at elevated temperatures17.
Experiment on the effect of various heat treatments has also been conducted on potato peels. It was observed that
steaming increase the chlorogenate content, while baking and frying almost completely eliminate the chlorogenate
present there in. The chlorogenic acid in potato peel remains constant upon boiling, but it may actually increase

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Sukrasno / Procedia Chemistry 13 (2014) 57 62

since the chlorogenic acid presence in the boiling water was not analyzed. The above observations suggest that heat
treatment might be needed to enhance the content of secondary metabolites during the preparation of the crude drug,
especially polyphenolic compounds. Removal of oxygen from the surrounding material may be needed to inhibit the
activity of polyphenoloxidase.
Heat treatments have also been recommended as one of alternatives to preserve the quality fruits and vegetables
before storage. Carotenoids and anthocyanines in immature fruit increase upon storage, and the increase is higher at
elevated temperature up to 25 qC and no synthesis at 30 qC or above. Increase of flavonoid is still taken place at low
temperature in chilly insensitive fruits such as apple. The changes of these metabolites are associated with fruit
ripening. Heat treatment through hot water dipping, rinsing and brushing, and hot air treatment have been
recommended to retard anthocyanin, ascorbic acid and chlorophyll degradations and carotenoid synthesis in
vegetables and fruits18.
4. Drying of Crude Drug
Drying of crude drug is aimed at removing water from the plant material in order to prolong the shelf life,
effective storage and transportation, and dose or weight accuracy. Fresh material contains large quantity of water
that rapidly evaporates even at ambient temperature. The present of water will also facilitate the growth of
microorganism especially in poor aeration condition. Biochemical reaction is also actively taking place in fresh
material and the constituents may substantially different from when the organ still attached to the original plant.
Nature of each material will determine the method of drying. High moisture containing materials such as rosella
calyx and Anredera cordifolia leaves needs a rapid drying method. Anredera cordifolia leaves strongly retain water
present in the tissue; aeration at room temperature will lead to the degradation of the material. Rosella calyx, in
addition to high water content also contains vitamin C that might be degraded during slow drying. Therefore, rosella
calyx that is dried by aeration at room temperature fails to produce red color upon boiling. Red color solution will be
obtained from boiling rosella calyx that is dried rapidly under the sun light or an air oven. Sun light will also degrade
some secondary metabolites present in the crude drug especially those are easily oxidized.
In most documents, it is recommended that drying should be conducted at temperature not higher than 60 qC.
However, in the cases of phenolic compounds drying at 60 qC should be avoided since peroxidase activity is the
highest at this temperature. The flavonoid content of Cosmos caudatus is even higher when pre-heated at 100 qC
compared to 60 qC and 80 qC. Flavonoid content of Cosmos caudatus leaves increase by 31 % when dried at 40 qC
while at 60 qC decrease by 22 %8.
Drying experiments on apricots at 55 oC and 75 oC also showed the decrease of catechin, epicatechin, rutin and
quercetine-3-glucoside19. Drying apricots at 55 oC decrease more chlorogenic acid, neochlorogenic acid, and
catechin compared to at 75 oC. However, for epicatechin, rutin and quercetin 3-O-glucoside higher decrease were
observed on drying at 75 oC compared to 55 oC. The antioxidant activity is higher in apricort dried at 75 oC than at
55 oC and both higher than in fresh. Decrease in polyphenols and increase antioxidant activity could be due to
several factors, such as increase antioxidant power of polyphenols at intermediate state of oxidation or increase in
reducing sugar and formation of Mailard Reaction Product known to have a great antioxidant activity. Air drying
treatment also increases the antioxidant activity of brocolli. High temperature and short time drying processes
maximized the antioxidant activity that might be due to the increase release of compounds from the matrix,
hydrolytic phenomena oxidation resulting in oligomers with higher antioxidant activity than the native compunds 20.
Another report mentioned that semidrying of tomato at 42 oC is darker than fresh tomato and the total phenolics,
flavonoid (rutin), lycopene and ascorbic acid are all lower than in fresh, so the antioxidant activity. In this
experiment, the tomato was cut into quarter before drying at 42 oC for 18 hours in force air drier21. Total phenolic
content of a number plant material also declined when dried at 55 oC15. The decreased of flavonoid and polyphenol
when dried at temperature around 50 oC might be due to the activity of polyphenoloxidase 22 and this explanation has
also been proposed20, 19, 23, 24. The possibility of flavonoid decreased due to Mailard reaction is unlikely takes place
at 60 oC, in addition pre-heating at 80 oC and 100 oC produced Cosmos leaves with higher level of flavonoids
compared to that of 60 qC8, 25, 26.

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Volatile oil content is mostly sensitive to drying and consideration must be taken in choosing the material in order
to produce the volatile oil. Most volatile oil from flower is best obtained from fresh flower. Similar cases were
observed with the oil from leaves. Ginger will only retain 17.7 % its oil content when it is sliced and dried.
Curcuma oil does not significantly different whether it is isolated from fresh or dry material. Crude drug containing
high boiling point oil component may maintain its oil content during drying. Vetiver oil is isolated through steam
distillation from partly dried material. The scent of dried vetiver root is much stronger compared to the fresh root.
Pogostemon oil is also mostly obtained from dry leaves. Table 1 showed the change of volatile oil yield when the
material producing the oil is dried.
Table 1. Comparison of Oil Content in Fresh and Dried in Some Crude Drugs
Crude Drug
Curcumae rhizome
Zingiberis rhizome
Piper betle leaves
Piper cf fragile leaves

Fresh (mL/1000 g)
1.19
8.40
2.40
3.80

Dried (mL/1000 g of fresh)


1.08
1.40
0.73
2.11

Loss of oil (%)


9.24
83.33
69.58
44.34

5. Conclussion
Slicing or cutting is necessary steps for crude drug preparation especially from big size plant organs or from
which the moist is hardly possible to be removed from the bulky organ. Slicing will expose part of the secondary
metabolites to the light and air. Sensitive metabolites may be oxidized and polymerized after cutting and drying.
This may explain the exhaustively extracted Curcuma rhizome with solvent that can dissolve curcuminoid well still
leaving residue with yellow color. While the extraction of fresh Curcuma rhizome with ethanol or methanol can
produce colorless residue. Eleutherina americana bulbs contain anthocyanins that can be completely extracted from
fresh bulb with hot water. However, extraction with hot water on sliced dried bulb or its powder still leaving reddish
residue. Thickness of the slices needs to be considered to minimize decomposition of the desired metabolites.
Intact garlic will contain allicin, after slicing alliin will be decomposed into allicin that give specific odor of
garlic. Similar observation can be made with onion or other Liliaceae species. This show substantial change takes
place during slicing. Substrate and the decomposing enzyme are both available in the organ, but the might be
separated in different compartments interact each other after the destruction of cell compartments during slicing or
grinding of fresh tissues.
Grinding is frequently conducted to enhance the efficiency of extraction process by increasing broken cells and
enhance the surface area that contact with the solvent. Caraway oil is much more effective to be extracted by super
critic fluid from ground than wholly seeds27. However, grinding may influence the content of the desired
metabolites. Volatile oil content of certain crude drug decrease upon grinding. This is partly due to the breaking of
oil glands that lead to the evaporation low boiling point component of the oil. Heat of the machine is also
contributing for the loss of oil from the crude drug upon grinding.
Table 2 Volatile oil content of crude drug before and after grinding
Crude drug

Original (ml/100 g)

After grinding (ml/100 g)

Loss (%)

Black pepper
Pimento
Cardamom with peel
Mace
Clove

3.37
3.19
3.61
16.10
17.30

2.21
2.71
2.20
9.10
11.5

34.42
15.05
39.06
43.48
33.53

Loss of volatile oil upon grinding can be minimized by cold grinding. Provision of chilled water and liquid
nitrogen jacket surrounding the grinding machine retain oil content of black pepper at 3.56 mL/100g and 3.60

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Sukrasno / Procedia Chemistry 13 (2014) 57 62

mL/100g respectively28.
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