Coffee, bean, roasted, ext.

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

publication

Title:

Acute toxicity of the aqueous extract of roasted and ground beans of Coffea canephora robusta in the Wistar rat

Author:

AKA Francis Beranger Angelo, Amonkan Kouao Augustin, Kahou Bi Gohi Parfait and NENE BI Semi Anthelme

Year:

2016

Bibliographic source:

The Pharma Innovation Journal 2016; 5(12): 01-05

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

not specified

GLP compliance:

not specified

Remarks:

No further information provided.

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

not applicable - High molecular weight constituents (Mwt > 719 g mol-1) such as polysaccharides, proteins and melanoidin polymers

Molecular formula:

not applicable - High molecular weight constituents (Mwt > 719 g mol-1) such as polysaccharides, proteins and melanoidin polymers

IUPAC Name:

not applicable - High molecular weight constituents (Mwt > 719 g mol-1) such as polysaccharides, proteins and melanoidin polymers

Constituent 2

Reference substance name:

Caffeine

EC Number:

200-362-1

EC Name:

Caffeine

Cas Number:

58-08-2

Molecular formula:

C8H10N4O2

IUPAC Name:

1,3,7-trimethyl-3,7-dihydro-1H-purine-2,6-dione

Test material form:

liquid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: roasted and ground beans of Coffea canephora robusta
- Expiration date of the lot/batch: not available
- Purity test date: not available

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: refrigerated
- Stability under test conditions: not available
- Solubility and stability of the test substance in the solvent/vehicle: not available
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not available

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The aqueous extract was obtained by infusion of roasted and ground beans of Coffea canephora robusta (Ccr). A filter coffee machine of Philips brand Daily collection insulated stainless timer HD7479/20, was used to prepare coffee. The infusion was made with 30 g of roast and ground beans of Ccr in 175 ml of distilled water. The filtrate obtained was evaporated in an oven at a temperature of 60 °C. The crystals obtained were pulverized. The captured fine powder was kept refrigerated in sterile glass jars sealed. This technique yielded 3g of dry extract.
- Preliminary purification step (if any): not available
- Final dilution of a dissolved solid, stock liquid or gel:
- Final preparation of a solid: not applicable

OTHER SPECIFICS:
To extract caffeine, one gram of freeze-dried coffee ground powder was transferred to a volumetric flask of 50 ml. An amount of 35 ml of methanol was added and the resulting suspension was immersed in an ultrasonic bath for 10 minutes. The suspension was subsequently cooled to room temperature and filtered on Whatman paper N°4.
The HPLC system consisting of a pump, a UV detector, and controlled by a computer (software) was used for assaying the proportion of caffeine in the extract. Qualitative analysis of caffeine was obtained by comparison of retentions times of the compounds eluted to the retention times of the reference solutions. The concentrations were determined from the average of the peak areas of the reference solutions. Caffeine content being the average of 3 tests, the concentration of caffeine in the extract is given by the following formula:
SC = S area x CW/ W area, where

SC = sample concentration
CW = Concentration witness
W area = peak area witness
S area = peak area sample

Test animals

Species:

rat

Strain:

Wistar

Remarks:

albino

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Pasteur Institute Adipodoumé, Abidjan, Ivory Coast
- Females (if applicable) nulliparous and non-pregnant: not specified
- Age at study initiation: 8-10 weeks
- Weight at study initiation: between 100 and 120 g
- Fasting period before study: not specified
- Housing: wire cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24 ± 3ºC
- Humidity (%): 50%
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12/12 hours

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

Test solutions and control solution were administered by a stomach tube.

Doses:

2000 and 5000 mg/kg bw

No. of animals per sex per dose:

3 female rats

Control animals:

yes

Remarks:

control animals received distiled water

Details on study design:

- Duration of observation period following administration: 14 days

- Frequency of observations and weighing:
Clinical signs were observed at 30 minutes, 1, 4, 8 , 24 and 48 hours.
The weight of each animal was determined before administration and then every third day.

- Necropsy of survivors performed: yes

- Other examinations performed: clinical signs, body weight,organ weights, histopathology, other:
Organs such as the kidney, liver and heart were removed and weighed.
The biochemical parameters such urea, blood sugar, creatinine, transaminases ASAT and ALAT and hematological parameters such as leukocytes, erythrocytes and platelets counts, hemoglobin, hematocrit, MCV, MCHC were determined.

Statistics:

The statistical analysis was performed using the Graph Pad Prism 5 software. The analysis of variance ANOVA (One-way ANOVA) followed by the Tukey-Kramer test was used for comparison of results. The difference is considered statistically significant when P <0.05.

Results and discussion

Effect levels

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 - < 5 000 mg/kg bw

Based on:

test mat.

Mortality:

Two deaths occured at the dose of 5000 mg/kg bw after 24 and 48 hours. No deaths related to the extract administration were recorded at the dose of 2000 mg/kg bw.

Clinical signs:

other: During the first hour after administration of the extract, the rats from experimental batches seemed agitated. This excitation was attributed to caffeine, which increases, neuromuscular transmission and increases the neural excitability reducing the disch

Gross pathology:

No effects were observed on liver, heart and kidney in experimental and control animals.

Other findings:

Caffeine content in the aqueous extract of roasted and ground beans of Coffea Canephora robusta was determined at 7.5%.

Any other information on results incl. tables

Effects on the organs weigh in animals receiving 2000 mg/kg of bw of the extract.

Organ (g)	Control rats	Experimental rats
Liver	4.08±0.41	4.17±0.28
Heart	0.43±0.26	0.47±0.14
Kidney	0.64±0.025	0.67±0.02

 

Effects on biochemical parameters in animals receiving 2000 mg/kg of bw of the extract.

Parameter	Control rats	Experimental rats
Urea (g/L)	0.19±0.01	0.18±0.02
Blood sugar (g/L)	0.95±0.16	1.05±0.20
Creatinine (mg/L)	8.67±0.95	7.67±1.04
Transaminase ASAT (U/L)	35.33±3.06	32.0±4.57
Transaminase ALAT (U/L)	39.33±1.67	37.67±2.62

 

Effects on leukocytes parameters in animals receiving 2000 mg/kg of bw of the extract.

Parameter	Control rats	Experimental rats
Polymorphonuclear neutrophils (%)	14.0±2.65	12.67±1.15
Polymorphonuclear eosinophils (%)	1.67±0.58	1.33±0.58
Polymorphonuclear basophils (%)	0.0±0.00	0.0±0.00
Lymphocytes (%)	79.33±3.08	81.0±2.00
Monocytes (%)	5.0±1.00	5.0±1.00

 

Effects on hematological parameters in animals receiving 2000 mg/kg of bw of the extract.

Parameter	Control rats	Experimental rats
Leukocytes (10^9/L)	8.43±2.21	10.53±1.77
Erythrocytes (10^12/L)	6.93±0.63	7.17±0.24
Hemoglobin (g/dL)	13.07±0.57	13.73±0.40
Hematocrit (%)	37.1±1.81	39.47±1.40
MCV (fl)	51.60±1.15	54.37±1.30
MCH (pg)	17.87±0.75	18.57±1.07
MCHC (g/dL)	34.40±1.02	34.67±0.85
Platelets (10^9/L)	728±109	753±127

Administration of roasted and ground beans of Coffea canephora robusta extract did not have any effect on biochemical and hematological parameters. Normal values of urea and creatinine suggest that this extract did not alter the structure and renal functions. In addition, the levels of transaminases (ALAT) and (ASAT) were not altered during the experiment indicating that liver and to a lesser degree, muscles were not affected.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In an acute oral toxicity study in rats, conducted according to OECD test guideline 423, a LD50 value was determined between 2000 and 5000 mg/kg body weight for aqueous extract of roasted and ground beans of Coffea canephora robusta containing 7.5% of caffeine.

Executive summary:

The acute oral toxicity study in rats, conducted according to OECD test guideline 423 was performed to assess the acute toxicity potential of aqueous extract of roasted and ground beans of Coffea canephora robusta.

Two sets of 3 animals received Coffea canephora robusta extract at 2000 mg/kg body weight, another set of 3 animals received a dose of 5000 mg/kg body weight and a control group were administered distilled water, by stomach tube.

Clinical signs including diarrhea, lethargy, excitability and death were observed at 30 minutes, 1, 4, 8, 24 and 48 hours. The relative weights of organs (kidneys, liver and heart) were recorded at the beginning of the study, and then every third day. At the end of the study, animals were sacrificed and gross pathology was performed. Additionally, the biochemical and hematological parameters were determined. 

No deaths were observed at 2000 mg/kg body weight, two deaths occurred at the dose of 5000 mg/kg body weight after 24 and 48 hours. There were no significant effects on the body weight in animals receiving coffee extract in comparison to control animals. Additionally, all animals from control and treatment groups gained weight during the experiment.

The administration of aqueous extract of roasted and ground beans of Coffea Canephora robusta did not have any significant effect on the relative weights of organs and biochemical and hematological parameters

The value of the lethal dose 50 (LD50) in this study has been determined between 2000 and 5000 mg/kg body weight, indicating that the aqueous extract of roasted and ground beans of Coffea Canephora robusta containing 7.5% of caffeine has relatively low acute toxicity and does not meet GHS criteria for acute toxicity classification.