An in vitro and in vivo investigation of the anti-inflammatory
properties of coal derived humates.
E.J. van Rensburg, G. Joonй and A.D. Cromarty, Department of Pharmacology, Faculty of Health Sciences, University of Pretoria, South Afr
INTRODUCTION:
mic substances occur widely in nature.
e therapeutic properties of humates have
escribed as antibacterial, antitoxic, anti-
genic, anti-arthritic, anti-allergic,
omodulatory and anti-inflammatory.
ious in vivo studies have been done to
strate the anti-inflammatory activity of crude
ts prepared from humus matter such as peat,
and sapropel.
documentation could be found concerning the
of coal-derived humates on inflammatory
ns. These preparations, especially products
d from brown coals, contain much higher
of high quality humate, are easier to produce
controlled conditions and are more available
re.
as been documented that agents that block
hesion molecule, CR3, expressed on the
e of activated neutrophils, are beneficial in the
ent of inflammation by inhibiting recruitment of
ytes into tissues.
AIM OF STUDY:
ermine:
ether oral treatment with potassium humate
period of a week could decrease the
matory effects caused by a contact
ensitivity reaction in vivo and to compare the
with prednisilone.
effects humates have on the expression of
y activated neutrophils.
RESULTS: Measurement of CR3 expression:
Contact hypersensitivity  Both potasium humate products caused a dose related
 Both the prednisilone and the leonardite derived humate product inhibition of CR3 expression by stimulated, but not resting
caused a significant (P < 0.5) decrease in ear swelling at 3h as well as neutrophils that was significant (p< 0.05) at 10µg/ml and higher
24 and 48h (Figure 2). Prednisilone proved to be superior to the (Figure 4).
leonardite humate whereas the humate product derived from
bituminous coal had no significant effect on ear swelling. Figure 4
 No signs of toxicity were observed during the 7 days of treatment The effects of a 15 min treatment with various concentrations of
with the humate products. However, the rats on prednisilone lost bituminous coal derived and the leonardite derived humate
weight compared to the control group (Figure 3). products on the expression of CR3 on resting and PMA-stimulat
human neutrophils determined by flow cytometry.
Figure 1
Control rat after DNFB challenge.
Figure 2
Difference in ear thickness between left and right ears of DNFB
challenged rats either without treatment or after a daily treatment for
one week, administered by gavage, of one of the following; 61mg/Kg
bituminous humate; 61mg/Kg leonardite humate; 1mg/Kg prednisilone.
The three columns represent the differences in ear thickness at 3
hours, 24 hours and 48 hours post challenge. Significant differences
compared to untreated controls.
* p< 0.05; ** p< 0.01
Figure 3
 METHODS:
e:
umate products were dissolved in water and
ested.
e produced from brown coal (leonardite)
ntains 92% soluble humate and fulvate).
e synthesized from bituminous coal (contains
% soluble humate).
ct hypersensitivity:
s experiment was done at the University of
a Biomedical Research Centre.
female Sprague Dawley rats of 8 to 10 weeks,
into four groups of 15 rats each.
day 0, the rats were weighed and sensitized
nting the shaved abdomen with 400µl of a
olution of 2,4-dinitro-fluorobenzene (DNFB) in
e:olive oil (4:1) and placed on one of the
ng oral treatments: (I) water only (2) leonardite
e (61mg soluble humate /Kg) (3) bituminous
umate (61mg soluble humate /Kg) (4)
silone (1mg/Kg).
day 6, the rats were challenged on the right
application to upper surface of the ear, of
a 0.5% solution of DNFB in acetone:olive oil
Figure 1).
ee hours after challenge both ears were
red with an engineering caliper across the ear
stance of 3mm from the tip. Each ear was
red 3 times and the average thickness
d. The measurements of the ears were
ed after 24 and 48 hours.
The changes in the body mass of rats after one week of treatment with
61 mg /Kg/day bituminous coal derived and leonardite derived humate
products or 1 mg/Kg/day prednisilone compared to untreated controls.
* p< 0.05; ** p< 0.01
DISCUSSION:
 The antiinflammatory properties of oral potassium huma
derived from leonardite compared favourably with
prednisilone
 Both of the humate products inhibited CR3 expression on
activated neutrophils.
 The difference in the activities between the two humate
products in the contact hypersensitivity model needs to be furthe
investigated.
 An over-expression of CR3 is associated with the production
a multitude of cytokines, reactive oxygen, nitrogen intermediates
and proteolytic enzymes that can cause tissue injury and lead to
inflammatory conditions.
 No signs of toxicity was observed with the two humate
products.
 Potassium humate has been proven to be safe in human
whereas prednisilone is associated with serious side effect
CONCLUSION:
The identification of a relative nontoxic
compound such as potassium humate tha
exerts its anti-inflammatory properties via th
blocking of an adhesion molecule that play
an key role in inflammation, is therefore an
exciting finding and merits further evaluatio
in the treatment of patients suffering from
inflammatory conditions.

rement of CR3 expression:

uspension of neutrophils was treated with
s concentrations of the humate products for 15
37°C, stimulated with 100ng/ml PMA and
ted for a further 15 min. CR3 quantitation was
y flow cytometry analysis on a Coulter Epics
C flow cytometer using an anti-CD11b FITC
lonal antibody (Beckman Coulter, Pallo Alto,
nia).

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