Forage Yield , Chemical Composition and In Vitro Gas Production of Triticale ( X Triticosecale wittmack ) and Barley ( Hordeum vulgare ) asociated with Common Vetch ( Vicia sativa ) Preserved as Hay or Silage

Forage cereals are deficient in protein, but legumes are acceptable in protein quality; the association of both forages can increase their nutritional value and biomass production. The objectives were to evaluate forage yield, chemical composition, energy content and In Vitro gas production of varieties of triticale (UAEMex and Siglo XXI), barley (Doña Josefa) and its association with common vetch, harvested at 145 days (milky-dough stage); a portion was ensiled and the other was hayed. Data were analyzed by a completely randomized design, and by using non-orthogonal contrasts to compare means of barley against triticale, common vetch and unicrops vs. associations. There were differences (P < 0.001) in DM (dry matter) production, being higher triticale Siglo XXI and associated with common vetch (6.50 and 5.4 ton DM/ha respectively), the highest CP content (P < 0.001) was for common vetch (234 g/kg DM) and their associations (187±2 g/kg DM). NDF content showed no differences between varieties and their associations (P > 0.302). Doña Josefa Barley showed a higher content (P < 0.05) of TDN (65%), NEl 1.4, and NEg0.8 Mcal/kg DM in both methods of preservation. Total gas production (ml/g DM), was higher for the association of Doña Josefa barley to common vetch (P < 0.05) (127±6 ml gas/g DM) compared to the rest (117±3 ml gas/g DM). As a conclusion, associations of barley or triticale to Siglo XXI with common vetch as hay or silage, allows us to consider them as an option to incorporate into the processes of production of livestock feed in high valleys.


Introduction
Forages are an important resource for animal feed in the world because they provide energy and protein for livestock (Ghanbari, 2000).Due to adaptation of highlands (located in altitudes of 2200 to 2600 meters above sea level), where the productivity of other forage is limited by low temperatures, an alternative to forage production are small grains such as wheat (Triticum aestivum), barley (Hordeum vulgare), oats (Avena sativa L.) and triticale (X Triticosecale wittmack) (Anil et al., 1998;Lema et al., 2004;Ross, King, O'Donovan, & Spaner, 2004).They are commonly used in winter forage by grazing (Lema et al., 2004) or conserved silage or hay (Haj-Ayed, González, Caballero, & Alvir, 2000).
Intercropping is defined as the growth of two or more crop species simultaneously in the same field during the growing season (Ofori & Stern, 1987).These intercropping fields are important for the development of production systems that depend little on external inputs (Adesogan et al., 2002).This may be due to the potential benefits of intercropping such as high productivity and profitability (Yildirim & Guvence, 2005); improved soil fertility by added nitrogen-fixing legumes (Hauggaard-Nielsen et al., 2001); efficient use of resources; and reduced damage caused by pests, diseases, and weeds (Banik et al., 2006).But their most important use is for the production of quality forage through the complementary effects of two or more crops simultaneously in the same area (Ross et al., 2004;Bingol et al., 2007;Lithourgidis et al., 2007).
Forage conservation from hay or silage; maintain the nutritional qualities for long periods of time (Givens & Rulquin, 2004;Scharenberg et al., 2007).In recent years, commercial varieties of triticale and barley have been released, counting on a wide range of advanced lines with specific adaptation to conditions in the highlands of Mexico.Such varieties are not available due to their potential as forage, nutritional quality or associations with common vetch, and use to establish feeding programs in ruminants.Except for some reports of forage yield and chemical composition, the technique of In Vitro gas production (Menke & Stengass, 1988;Theodorou, Williams, Dhanoa, Mc Allan, & France, 1994) is a parameter of importance to know fermentation patterns and degradation, depending on the availability of nutrients from the feed.The objectives of this study were to evaluate the forage yield, chemical composition and In Vitro gas production of two varieties of triticale and barley associated with common vetch as silage or hay.

Crop Establishment
The study took place in the valley of Toluca, Mexico State, located at coordinates 19° 17' north latitude and 99º 39' west longitudes at an altitude of 2675 meters above sea level.The climate is of type C (w2) (w) b (i'), corresponding with subhumid climate with summer rains, with little temperature variation between 5 and 7°C.Average annual rainfall is 1000 mm and mean annual temperature is 14°C.The predominant soil type is vertisol pelvic volcanic (INEGI, 2000).
The evaluation consisted in two varieties of spring triticale (UAEMex and Siglo XXI) and barley (Doña Josefa) as unicrop and associated with common vetch, which were provided by the Institute of Agricultural Research and Training in Mexico State (ICAMEX) in coordination with the International Maize and Wheat Improvement (CIMMYT) and the Faculty of Agricultural Sciences at the Autonomous University State of Mexico.The trial was established on April 24, 2010 (three replicates for each of the varieties).The experimental unit consisted in seven rows, each 50 linear meters by 30 cm wide (with two lines of planting and spaced at 80 cm), leaving a meter on each side to protect the experimental area.The planting density was 80,000 plants/ha, which was fertilized with 60-90-70 -N-P-K, respectively (130.4 kg of urea, 195.6 kg triple superphosphate, and 116.7 kg potassium chloride/ha), and irrigation was applied after sowing.

Sampling, Forage Conservation and Chemical Analysis
Each sample of dry matter (DM) and fresh matter (FM) was performed by cutting the crop at an approximate height of 2 cm from the soil surface, when the grain was in the milky-dough stage (145 days); random sampling for the collection of varieties, taking 1.0 m linear from areas of the central rows forming the experimental unit, obtaining three samples of each variety, and determininged the DM content (60°C for 48 h) and moisture for forage yield (ton/ha).
Of the selected forage samples, a part was hayed (three samples of each variety and association were sun-dried for seven days to reach approximately 85% DM), and the other part was prepared freshly chopped for micro-silages by triplicate, in 20 x 10 cm tubes of polyvinyl chloride, with a capacity of 1.5 kg (Cobos et al., 1997), and compacting and sealing them well with plastic bags and tape to prevent the entrance of air.At 60 days the samples were opened from micro-silages (200 g), dried in a forced air oven (60°C, 48 h), and ground in a Willey mill with a sieve of 2 mm.Forage samples were analyzed for DM (#934.01),ash (#942.05),and N (#954.01)according to (AOAC, 1997).The neutral detergent fiber (NDFom, Van Soest et al., 1991), acid detergent fiber (ADFom), and lignin (AOAC, 1997;#973.18)analyses used an ANKOM200 Fiber Analyzer Unit (ANKOM Technology Corporation, Macedon, NY, USA).NDFom was assayed without use of an alpha amylase but with sodium sulfite in the NDFom.Both NDFom and ADFom are expressed without residual ash.Moisture content of the silages was determined through distillation with toluene (Haigh & Hopkins, 1977).

In Vitro Gas Production
For the in vitro gas production technique, we used three rumen fistulated dairy cattle (LW 450 ± 20 kg) as donors of rumen fluid.The animals received a diet of oat hay and alfalfa hay (50:50), formulated to meet all of their nutrient requirements (NRC, 2001), and was supplied twice a day (8:00 and 16:00 hours).Fresh drinking water was available to the cows at all times.
Gas production was determined in 125 ml amber flask and three series of incubation for each sample of forage conservation method, using the technique proposed by Theodorou et al. (1994).In each flask was introduced 0.8g DM of each of the samples, then we added 90 ml of buffer solution (Menke & Steingass, 1998) gassed with CO 2 and stored (4°C for 12 h); until the next day when we took 700 ml of ruminal fluid and 300g of solid rumen contents of each cattle donor, and filtered the homogenized mixture through four layers of gauze and glass wool subsequently, maintaining the rumen fluid at 39°C, then was gassed with CO 2 , and subsequently added to each flask 10 ml of ruminal fluid.Finally, the flasks were introduced into a water bath at 39°C and initiated gas production record using a pressure transducer (DELTA OHM, Manometer, 8804).The volume of gas produced was recorded at 3,6,9,12,24,36,48,72 and 96 h of incubation.For corrections were used two flasks without substrate as blank and barley straw as standard.

Calculations and Statistical Analysis
After the incubation period (96 h), the accumulated gas was released and the fermentation residues of each flask were dried (60°C, 48 h) to calculate the proportion of dry mater disappeared (DMd) and relative gas production (RGP, ml gas g DMd) according to Gonzalez Ronquillo et al. (1998).
The kinetics of gas production were determined by adjusting the model, (1) Proposed by Krishnamoorthy et al. (1991); according to the model, b represents the total production of gas (ml gas/g initial DM); c the rate of degradation in relation to time (h -1 ), and t time (h -1 ).
The values of net energy for lactation (NE l ) and net energy gain (NE g ) of forages were obtained through determining the content of total digestible nutrients (TDN) from chemical composition (NRC, 2001): Total digestible nutrients (TDN) Where Were Y is the variable, Txi is the effect duo to the treatment, and εij is the experimental error.
We used non-orthogonal contrasts to compare: C1, barley against triticale; C2, barley-common vetch vs. triticale associated with common vetch; C3, common vetch vs. barley and triticale associated with common vetch; C4, unicrops (barley and triticale) vs. barley and triticale associated with common vetch.The comparison of measures of each variable was carried out through the Tukey method (Steel et al., 1997).

Forage Yield, Chemical Composition and Energy Content
Table 1 shows the forage yield (ton/ha) for the varieties of barley and triticale, as well as its association with common vetch; differences were observed (P < 0.001) between forages, being higher (C2, P < 0.020), common vetch when compared to their associations (C3, P < 0.013), when comparing forage associations was higher (C4, P < 0.001) than unicrops.Except triticale, S XXI was higher than barley Doña Josefa and triticale UAMex.
Table 1 shows forages conserved as hay; CP content was higher for barley (C1, P < 0.009) vs. triticale's, and its association with common vetch (C2, P < 0.005); when comparing C3 (P < 0.001), common vetch was higher with respect to their association; C4 with the unicrops were lower (P < 0.001) with respect to associations with common vetch.NDF content were not different between forage varieties and their associations (P < 0.302), but ADF content was higher (P < 0.001) for triticale's vs. barley, and its association with common vetch (C2, P < 0.05).Likewise, common vetch was lower when compared with their associations (C3, P < 0.019); associations with common vetch were higher (C4, P < 0.001) compared with unicrops.There were differences for TDN content, NE l and NE g (P < 0.001) between varieties and their associations (P < 0.001).NE l and NE g were higher (C1, P < 0.001) for barley compared with triticale's and their association with common vetch (C2, P < 0.02).
Common vetch was higher in terms of TDN content, NE l, and NE g (C3, P < 0.005) when compared with their associations.Table 2 presents the chemical composition of forage silage; CP content was higher (C1, P < 0.010) for barley vs. triticales.Common vetch (C3, P < 0.007) was higher with respect to their association; when compared to unicrops it was lower (C4, P < 0.001) compared with their associations.There were no differences for C1 (P < 0.304) and C2 (P < 0.104); NDF content was higher for varieties associated with common vetch (C3, P < 0.025), the association barley and triticales associated with common vetch was higher (C4, P < 0.022) compared with the unicrops.ADF content was no different for C1 (P < 0.187) and C4 (P > 0.614), ADF content was higher (C2, P < 0.023) for triticale compared with barley associated with common vetch; common vetch was higher (C3, P < 0.015) when compared with their associations.TDN, NE g and NE l show differences (P < 0.001) between varieties and their associations.TDN, NE l , and NE g were higher (C1, P < 0.001) for barley compared to triticale, and their association with common vetch (C2, P < 0.02).Common vetch was lower (C3, P < 0.015) compared with their associations with barley and triticale's, TDN, NE l and NE g content were higher for associations (C4, P < 0.001) compared with unicrops.

In Vitro Gas Production
There were differences (P < 0.001) for the total gas production (b) for the different forages evaluated as hay (Table 3); barley was higher (C1, P < 0.001) compared with triticale's (Figure 1), and there was a trend (P < 0.092) when compared with common vetch-barley vs. common vetch-triticales.Common vetch was lower (C3, P < 0.003) when compared with their associations (Figure 2), and unicrops were lower (C4, P < 0.001) compared with their association.There were no differences for the fermentation rate for C2 (P < 0.116) and C3 (P < 0.433).
Fermentation rate (c), shows differences (P < 0.001) between treatments, being higher (C1, P < 0.001) triticale's compared with barley; and associations were higher (C4, P < 0.004) respect to unicrops.There were no differences for lag time in C2 (P < 0.117); lag time in barley was higher (C1, P < 0.001) compared with triticale, and common vetch was higher (C3, P < 0.001) compared with their associations.Unicrops were lower (C4, P < 0.028) compared with their associations.There were no differences (P > 0.1) for DMd between forages and their associations.Significant differences were observed (P < 0.003) in RGP, being higher in barley (C1, P < 0.044) compared with triticale,, and associated with common vetch (C2, P < 0.022).Common vetch was lower (C3, P < 0.008) compared with their associations, and common vetch associated with barley and triticale's was higher (C4, P < 0.006) compared with unicrops..006b = total gas production (ml gas/g DM initial), c = degradation rate vs. time (h), lag time (h); DMd = dry matter disappeared at 96h (mg/100 mg ), RGP = relative gas production (ml gas 96h/g DMd 96h).There were differences (P < 0.001) for the total gas production (b) for the different forages evaluated as silage (Table 3); barley was higher (C1, P < 0.001) compared with triticale (Figure 3), and there was a trend (P < 0.092) when compared common vetch-barley vs. common vetch-triticale.Common vetch was lower (C3, P < 0.001) when compared with their associations (Figure 4), and unicrops were lower (C4, P < 0.001) compared with their   Carr et al. (2004) indicates that the cultivation of barley and oats associated with pea (Pisum sativum) increased forage yield (0.63 and 0.72 ton/ha) compared with unicrops.However, Gonzalez et al. (2007), evaluated genotypes of triticale, oats, and barley and observed a differential behavior in unicrops and their association with common vetch, since triticale had lower performance in unicrop and their association; however, oats and barley increased their forage yields in association with common vetch, reaching total forage yields of 4.80 and 3.99 ton/ha.Fernandez et al. ( 2007) associated triticale with pea, with yields of 4.8 and 6.6 ton/ha, coinciding with the results of the present study.This shows an effect on the species, considering that the cereals were harvested when the grain was in the milky-dough stage and common vetch at the end of the flowering stage; the amount of forage yield was higher compared to that obtained by associating common vetch with triticale and barley.This coincides with Lauriault et al. (2002) where common vetch had similar forage yield to that observed in the present study; and similarly by associating common vetch with cereals, this legume crop provided the least amount of DM to the total amount of forage produced, due to the effect of competition by cereals over legumes.Anil et al. (1998) founded that when associating cereal with a legume forage, their quality gets better; these results agree with other studies (Khorasani et al., 1997;Carr, Horsley, & Poland, 2004) conducted in temperate zones, and similar to those in North America.Cereals in general are low in protein, however, the values of protein may reach 19% prior to ear emergence for barley, diminishing rapidly to plant maturity (Hargreaves, 1994), but this can vary depending on the species, variety and weather conditions.The present study highlights the barley Doña Josefa associated with common vetch with a higher protein content; similar data have been reported in other studies that have included a legume associated with forage cereals, and found that in these systems significantly increased the CP content in barley as the present study (Caballero, & García, 1996;Haj-Ayed et al., 2000;Assefa & Ledin, 2001;Kuusela, 2004;Carr et al., 2004).

Barley
NDFom and ADFom content increase in barley and triticale compared with their associations.Contrary to Ross et al. (2004), showing an effect of the legume on the reduction in NDF content in intercrops than in unicrops (Ghanbari & Lee, 2002), so the benefits of the association reflected an increased intake and digestibility of forage.Garduño et al. (2008), studied oats in association with common vetch and founded lower values in NDF! compared to cereals evaluated in the present study, but similar in ADF.Lignin content for intercropping increased, probably due to the contribution of common vetch.Ghanbari-By Lee (2002) and Haj-Ayed et al. (2000) report that in association with legumes, the lignin content increased; this is because those legumes have lower cell wall content and higher lignin content (Parra et al., 1972).The nutritional value of associated varieties is directly related to the cutting age, showing a positive response in associated systems (intercrops).Combining the growth of forage-grain crops (cereals) with crops that are able to increase the protein content of them as legumes, it is possible to obtain a higher forage production and acceptable quality (Ghanbari, 2000).
The percentage of TDN was different (P < 0.01) for all forages tested, with values ranging from 59 to 67%, in both methods of conservation; similar results were reported by Carr et al. (2004), who evaluated barley and oats associated with peas, reporting values of 51 and 57% TDN.NE l and NE g values were differents for the forages evaluated, highlighting barley Doña Josefa and their association with common vetch, with the highest energy content with respect to triticale and their associations in both methods of preservation.This coincides with Gonzalez et al. (2007), who highlight barley and their association with common vetch, with higher averages in both (1.2 NE l and 0.5 NE g ).Quiroz (2006), report a higher energy content of triticale compared to other commercial varieties of triticale and oats; Elizalde & Gallardo (2003), found a lower NE value for barley, contrary to the present study.

In Vitro Gas Production
Lara Fuentes & Valdez Arciniega (2010), founded values ranged from 305 and 275 ml gas/g DM in barley varieties with different levels of N fertilization.Moreover, evaluating different varieties of whole corn plant, Antolin et al. (2009) reported mean values of 217 and 202 ml gas/g DM for silage and hay, respectively.This indicates that varieties of triticale and barley fermented less compared to whole-plant corn and other barley varieties.Catrileo et al. (2003) evaluated the in vitro degradation of barley with clover (Trifolium repens and Trifolium pratense) silage, noting a decrease with the increasing maturity of the association, but there was not an influence of the species that were associated with barley.This is contrary to the present study, which indicates that common vetch had a positive influence on gas production of the cereals evaluated.

Conclusions
Both barley and triticale associated with common vetch showed a higher fermentation compared with our association; considering its nutritional quality and energy content, forage cereals associations with common vetch is an option with higher forage yield and improved nutritional content for feeding livestock, both preserved as hay or silage.

Table 3
. Parameters of In Vitro gas production (ml gas/g DM) obtained from the adjustment of incubation and digestibility of barley, triticale, and common vetch and their associations, preserved by hay