Impact of geranium essential oil as a sustainable feed additive on in vitro rumen gas and methane production, nutrient degradability and fermentation efficiency

	CFM¹	Berseem hay	Rice straw	Diet²
Dry matter	903	890	940	902
Organic matter	923	884	851	900
Crude protein	165	128	42	138
Ether extract	47	54	19	47
Nonfibrous carbohydrates	414	224	166	313
Neutral detergent fiber	297	478	624	402
Acid detergent fiber	175	381	394	279
Metabolizable energy (Mcal/kg DM)³	3.60	2.65	1.98	3.06

1 Concentrate feed mixture (CFM) contained per kg DM: 170 g soybean meal, 395 g wheat bran, 395 g maize, 20 g limestone, 10 g vitamins and minerals mixture, and 10 g salt.

2 Diets: Contained per kg DM: 500 g concentrate mixture, 400 g berseem hay and 100 g rice straw.

3 Calculated (NRC, 2001).

Table 2.

Volatile compounds in geranium oil (Pelargonium graveolens L.) identified by GC-MS analysis

Compound¹	RT²	Concentration³ (%)
α-Pinene	5.19	1.1
Myrcene	7.94	0.5
Linalool	10.18	7.9
Menthone	10.99	2.0
Isomenthone	12.89	8.3
α-Terpineol	13.86	1.1
Citronellol	15.41	32.4
Trans-Geraniol	16.55	13.3
Citronellyl formate	17.21	15.4
Geranyl formate	18.39	3.4
β-Caryophyllene	23.20	1.8
Isoledene	24.07	4.0
Muurolene	25.56	1.7
Guaia-6,9-diene	27.29	0.8
Phenylethyltiglate	29.62	1.4
Eudesmol	30.84	4.9

1 Identification based on authentic standards, National Institute of Standards and Technology (NIST) library spectra, and literature.

2 RT, retention time (min).

3 Concentration based on the total areas of the identified peaks.

*In vitro* ruminal gas production (mL/g incubated DM) of a total mixed ration supplemented with different levels of geranium oil

Table 3.

In vitro rumen gas production (GP), methane (CH₄) and carbon dioxide (CO₂) kinetics as affected by increasing levels of geranium oil (%, dry matter basis)

Level	A	C	Lag	A	c	Lag	%⁴	A	c	Lag	%⁴
0	78^c	0.076^a	1.55	28.6^b	0.035	1.42	29.3^a	58.8^c	0.036^c	2.47	63.5
0.5	108^b	0.084^a	1.62	35.6^b	0.036	1.56	27.5^ab	75.0^b	0.045^b	2.37	62.5
1	116^b	0.078^a	1.56	36.5^b	0.042	1.54	27.9^ab	79.1^b	0.048^ab	2.18	62.7
1.5	132^a	0.060^b	1.65	38.9^a	0.035	1.56	25.4^b	88.3^a	0.053^a	2.35	65.5
SEM	2.0	0.0020	0.09	1.05	0.0037	0.061	0.77	1.65	0.0017	0.241	1.28
P value
Treatment	<0.001	0.001	0.852	0.007	0.519	0.356	0.045	<0.001	0.008	0.859	0.392
Linear	<0.001	0.563	0.962	0.007	0.212	0.193	0.231	<0.001	0.002	0.420	0.683
Quadratic	<0.001	0.044	0.425	0.002	0.460	0.228	0.025	<0.001	0.004	0.883	0.493

Means in the same column with different superscripts differ (p<0.05). The p-value represents the observed significance level of the F-test for the treatment effect; SEM stands for the standard error of the mean.

1 GP parameters: A is the asymptotic GP (ml/g DM), c is the rate of GP (/h), Lag is the initial delay before GP begins (h).

2 Methane (CH₄) production parameters: A is the asymptotic CH₄ production (ml/g DM), c is the rate of CH₄ production (/h), Lag is the initial delay before CH₄ production begins (h).

3 Carbon dioxide (CO₂) production parameters: A is the asymptotic CO₂production (ml/g DM), c is the rate of CO₂ production (/h), Lag is the initial delay before CO₂ production begins (h).

4 The proportion at the end of incubation (48 h).

*In vitro* ruminal methane (CH₄) production (mL/g incubated DM) of a total mixed ration supplemented with different levels of geranium oil

*In vitro* ruminal carbon dioxide (CO₂) production (mL/g incubated DM) of a total mixed ration supplemented with different levels of geranium oil

Table 4.

In vitro rumen fermentation profile of diet with increasing levels of geranium oil (%, dry matter basis)

Level	dDM	dNDF	dADF	Total	C2	C2 (%)	C3	C2 (%)	C2: C3	C4	C2 (%)	pH	NH3-N	ME	PF24	MCP	GY24
0	473^c	439^b	412^c	23.4^c	11.4^c	48.9	7.90^b	33.7	1.47	4.08	17.4	6.27^b	10.4	4.68^b	6.22^a	328	139^b
0.5	511^bc	489^ab	463bc	27.4^b	13.7b	50.0	9.25^a	33.8	1.48	4.42	16.1	6.60^a	10.6	5.31^a	4.83^b	306	183^a
1	536^ab	510^a	492ab	27.3^b	13.9b	51.1	9.80^a	35.9	1.42	3.54	12.9	6.53^a	10.7	5.44^a	4.74^b	321	183^a
1.5	574a	539^a	524a	30.5^a	15.8a	52.0	9.80^a	32.1	1.62	4.84	15.8	6.50^ab	10.8	5.50^a	4.63^b	354	175^a
SEM	11.0	12.0	12.3	0.34	0.40	2.00	0.292	1.21	0.100	0.517	1.73	0.055	0.28	0.046	0.151	12.5	5.3
P value
Treatment	0.001	0.002	0.001	<0.001	0.004	0.716	0.005	0.258	0.577	0.395	0.376	0.013	0.804	<0.001	0.002	0.126	0.001
Linear	0.003	0.003	0.002	<0.001	0.002	0.446	0.002	0.239	0.732	0.487	0.106	0.009	0.468	<0.001	0.001	0.679	0.004
Quadratic	0.008	0.011	0.010	<0.001	0.008	0.624	0.049	0.168	0.345	0.151	0.649	0.027	0.645	<0.001	0.001	0.689	0.009

1 dDM is dry matter degradability (mg/g incubated), dNDF is neutral detergent fiber degradability (mg/g incubated), dADF is acid detergent fiber degradability (mg/g incubated).

2 SCFAs is short chain fatty acids (mmol/L), C₂ is acetate (mmol/L), C₃ is propionate (mmol/L), C₄ is butyrate (mmol/L).

{ label (or @symbol) needed for fn[@id='j_aoas-2025-0118_tfn_015'] } ₃NH₃-N is ammonia-N (mg/g DM), GY₂₄ is gas yield at 24 h (mL gas/g DMD), ME is metabolizable energy (MJ/kg DM), PF₂₄ is the partitioning factor at 24 h of incubation (mg degradable DM: mL gas), MCP is microbial CP production (mg/g DM).

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DOI: https://doi.org/10.2478/aoas-2025-0118 | Journal eISSN: 2300-8733 (formerly 1642-3402) | Journal ISSN: 1642-3402

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Language: English

Submitted on: Apr 1, 2025

Accepted on: Oct 20, 2025

Published on: Feb 16, 2026

Published by: National Research Institute of Animal Production

In partnership with: Paradigm Publishing Services

Publication frequency: 4 issues per year

Keywords:

degradability,

geranium essential oil,

in vitro fermentation,

methane,

phytogenics

Related subjects:

Medicine,

Veterinary medicine

© 2026 Ahmed E. Kholif, Moyòsore J. Adegbeye, Gouda A. Gouda, Tarek A. Morsy, Einar Vargas-Bello-Pérez, published by National Research Institute of Animal Production
This work is licensed under the Creative Commons Attribution 3.0 License.

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Impact of geranium essential oil as a sustainable feed additive on in vitro rumen gas and methane production, nutrient degradability and fermentation efficiency

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