Effect of Peppermint and Rosemary Extractions on Ruminant In- vitro Digestibility

| The objective of this study was to evaluate the effects of organic, watery and alcoholic extractions of peppermint and rosemary leaves on in-vitro ruminant digestibility. Three tubes for each treatment with three blank were used and the mean value was evaluated statistically. It was found that after 48 hours of incubations, addition of 200mg of organic extractions for peppermint and rosemary significantly decreased dry matter digestibility (DMD) in contrast with control, by 83.41%, 81.99% and 86.21% respectively, and organic matter digestibility (OMD), 85.23%, 84.27% and 90.19% respectively, while there were no significances for adding 200mg of dried leaves, watery and alcoholic extractions. Results suggested that the addition of peppermint or rosemary as a dried herbage or fresh to ruminants’ diet didn’t alter the rumen digestibility with a high content of crude protein.


Advances in Animal and Veterinary Sciences
October 2019 | Volume 7 | Issue 10 | Page 911

PreParaTiOn Of PlanT exTracTs
The leaves and flowering plants of peppermint and rosemary were sampled and dried at 65°C in a ventilated oven for 48 -96h. then grounded at sieving 1mm. , stems and woody parts were separated. Three methods of extractions, watery, alcoholic and organic were used at room temperature (Shtayeh and Abu Ghadeib, 1999;. Watery extracts, 50gm of samples were extracted in a 500ml beaker, fill it with boiled distill water with stirrer for 30 minutes then filtered with cheese cloth and dried at 40°C. Alcoholic extracts: 100g of peppermint or rosemary was extracted with 500 ml of 70% ethanol in a 1000ml beaker with stirrer for 24h ., then filtered with cheese cloth, toke the supernatant fluid and dried at 40°C to evaporate alcohol and dried. Organic extract: 100gm of samples were weighted in a 1000ml beaker, fill it with hexane and soak for 24h. then extracted with a Soxhlet apparatus during 2h. All extracts were stored in deep freeze until use. Nutritional value was analyzed according to AOAC (2012) ( Table 1).

deTerminaTiOn Of TOTal PHenOlic cOnTenT
Total phenolic were determined with Folin-Ciocalteu reagent. Gallic acid was used as a standard (we can use tannic acid, lignin or gallic acid), and the total phenolic were expressed photometrically as gallic acid equivalents to 100gm of raw material using Lambert-bear law "A = As×B×C" Where: A(absorption), As (0.966) from the calibration curve, B=1 (cell diameter) and the C (concentration of phenolic compounds (mg/ gm)). The Folin-Ciocalteu reagent reducing polyphenols and producing blue color which measured spectrophotometrically at 765nm (Cuvelier et al., 1996).

In-vItro digesTibiliTy
In-vitro digestibility was implemented as Tilley and Terry (1963): Active sample of rumen liquor was collected from slaughtered sheep and incubated in water bath at 38°C, artificial saliva prepared by two solutions, first: McDougall's Buffer solution, 49gm NaHCO3 + 18.6gm Na2H-PO4 dissolve in 800ml DW, second: Chlorides solution, 23.5gm NaCl + 6gm MgCl2.7H2O + 28.5gm KCl + 2gm CaCl2 dissolved in 1000ml DW. Add 100ml from the second solution to the first one (800ml) and completed up to 1000ml then incubated in 38°c water bath. Digestion tubes with 0.5gm grinded alfalfa hay through a 1mm sieve were prepared with two levels of peppermint or rosemary extracts (0, 200mg per tube). A mixture of 40ml artificial saliva and 10ml rumen liquor was added to each tube then incubated for 48h in shaker water bath at 38°c with gentle shaking two times a day. After finished 48h centrifuged the tubes and dry the residue (precipitate) 105°c overnight, weight then ash at 600°c for 5 h. calculate the dry matter and organic matter digestibility. All chemical compositions were determined according to AOAC (2012) for dry matter (DM), wet, organic matter (OM), crude protein (CP), ether extract (EE), crude fiber (CF), ash and nitrogen free extract (NFE) as % of DM basis.

sTaTisTical analysis
One-way ANOVA analysis was performed to determine significant differences (P<0.05) between treatments, which was implemented using statistical program (SAS, 2012) and Duncan test (Duncan, 1955 (Table 2), main effect of added extractions didn't differ significantly in contrast with control ( Figure 1), hexane is an organic solvent that is very efficient in dissolving fat and fat-like substances (such as vitamins and active soluble compounds), therefore, there is a possibility of increasing their concentration and affect negatively on in-vitro digestibility, the same changes of phytofactors availability to the microorganisms has been described by

Advances in Animal and Veterinary Sciences
October 2019 | Volume 7 | Issue 10 | Page 912 identify effective compounds able to positively affect rumen ecosystem, especially with in-vivo digestibility and continues flow rate of nutrients, water.

effecT Of PePPerminT Or rOsemary On In-vItro Organic maTTer digesTibiliTy
Results of organic matter digestibility (OMD) indicated to significant decreases (P<0.05) for in vitro digestibility with 200mg organic extraction of peppermint and rosemary in contrast with control, 85.23%, 84.27% and 90.19% respectively, but no statistical difference was observed between dried leaves, watery and alcoholic extractions (Table 3). Digestibility affects by organic matters content, incubation time, proportions of rumen microorganisms, the relatively higher OM degradability may be according to the low content of ash (Table 2) or due to the phytofactors in extracts which it non degraded by rumen microorganisms. On the other hand, no differences for the main effects of added extracts in contrasted with OM digestibility of alfalfa hay (Figure 1), Hosoda et al. (2006) showed a limited effect of peppermint on rumen digestibility in dairy cows, while Cobellis et al. (2016) demonstrated that rosemary leaves may be used to modulate rumen microbiome and its function to more utilize rumen nitrogen. Some of active component as short chain fatty acids work as antioxidant and have more than one mode of action, so, the strong antioxidant activity my appeared with organic extractions. Secondary metabolites in plant extracts contain antimicro-bial properties, but mode of action differs in extract rather than in plant, that may play the main role of non-significances for adding extract in contrasted with control (Figure 1) and increased apparent OM digestibility when fed 200mg/sheep/ day (Sahraei et al., 2014).

AcKnOwlEdgEMEntS
This study was supported by College of Agricultural Engineering Sciences, University of Baghdad. The authors are grateful to the dean of college and the head of Animal Production Department in addition to the head of Horticulture Department for their assistance during the experimental period.

cOnFlIctS OF IntErESt
Authors declare that there are no conflicts of interest.

AutHOrS cOntrIbutIOn
All authors contributed to the work, discussed the results and contributed to the final manuscript.