ANALYZING SILAGES FOR FERMENTATION END PRODUCTS

  Limin Kung, Jr., Ph.D.  
Dept. of Animal & Food Sciences  
University of Delaware  
Newark, DE 19717-1303  

and

  Martin R. Stokes, Ph.D.  
Dept. of Biosystems Science & Engineering  
University of Maine  
Orono, Maine 04469-5763  

 

Analyzing feeds for nutrient content is essential for proper balancing of dairy rations.  This is especially true for those feeds that have undergone the fermentation process.  Historically, silage samples have been subject to analyses that included determination of dry matter, protein fractions (degradable and undegradable protein, soluble protein, and heat damaged protein), ADF, NDF, mineral content and energy (calculated from ADF content).  In addition to these components, lactic, acetic, and butyric and propionic acids, pH, ammonia nitrogen, ethanol, and yeast and mold counts are quantified in research experiments.  Recently, some laboratories have offered a more complete nutrient analysis of silages that includes many, if not all, of these variables. What do these analyses mean?

 

First, the pH of silage measures the degree of acidity.  A pH of 7 is neutral and numbers less than 7 indicates acidic conditions.  A rapid production of acids inhibits bacteria while removal of air inhibits the growth of yeasts and molds.  The pH of silage is easy to determine.  A silage sample can be prepared by vigorously shaking a small sample of silage with 100 ml of water (about 4 fluid ounces) for a minute or two.  A pH-sensitive paper can be dipped into the liquid and will change color-based on the degree of acidity.  This can be compared to a color chart and converted to an approximate pH reading.  Producers can obtain pH-sensitive paper with a cost of a few cents per test.  A hand-held battery operated pH probe can give a more accurate measure of acidity and can be purchased for about $40-60 but must be calibrated before each use.  (Analytical labs determine pH using an even more accurate pH meter.)  Fresh alfalfa forage and corn forage have pH values of about 5.8 to 6.5 and 5.0 to 5.3, respectively.  Well preserved corn silage will have a pH in the range of 3.7 to 4.2 and the pH for well preserved high moisture corn will range between 4.0 and 4.5.  Because alfalfa has a high buffering-capacity, its silage pH will be slightly higher (4.5 to 5.0).  In alfalfa silage, pH is usually higher in high-dry matter content silage.  For example, 30% DM alfalfa silage may have a pH of 4.5 while 50% DM alfalfa silage may have a pH of 4.9 or higher.  In general, a low pH means that more acid was produced.  On the other hand, a high silage pH can have several causes.  First, it may mean that the silage did not ferment well.  Cold environmental conditions (late fall harvesting), lack of sufficient substrate for bacteria to make the acids, an undesirable fermentation (e.g. clostridia), or a low moisture content can each lead to a high pH.  A high pH could also be an indicator of silage that has spoiled due to exposure to air; badly deteriorated silages may have a pH greater than 7.5.

 

Lactic acid is the strongest and most abundant acid produced during an ideal fermentation.  Moderate levels of lactic acid are an indication of good fermentation.  The lactic acid content of good corn and alfalfa silage ranges from 4 to 6% and from 3 to 8%, respectively on a dry matter (DM) basis.  In high moisture corn, lactic acid is normally found in the 1 - 3% range.  Of the volatile fatty acids (VFA), acetic acid is found in the greatest concentration and butyric and propionic acids are uncommon end products of normal silage fermentation.  The acetic acid content of corn and alfalfa silages is usually between 1 to 3% and butyric acid content should be undetectable in good silages.  High moisture corn usually has a low acetic acid content (less than 1.0%).  High levels of acetic (> 3 to 4%) or butyric acid (> 0.5%) in any type of silage are indicators of less than desirable silage fermentation.  Ironically, such silage will probably be quite stable when exposed to air (even though the pH may be quite high in silages high in butyric acid).  Silages with high amounts of acetic and butyric acids have been associated with low dry matter intakes.  Moreover, silages high in butyric acid are usually low in energy and have undergone extensive protein degradation resulting in large increases in the soluble protein fraction and losses of dry matter.  A silage with high levels of butyric acid could have been caused by ensiling a high protein, low sugar forage, such as alfalfa, too wet (< 30% DM).  It can also be caused by clostridia from manure that was applied too close to the harvest date (allow 21 to 28 days if possible).

 

The ratio of lactic acid to acetic acid is a good indicator of the efficiency of the silage fermentation.  Lactobacilli are very efficient at producing lactic acid but acetic acid production also involves a loss of carbon from the silage during fermentation.  This is equivalent to a loss of DM from the silo and there will be less DM available in high acetate silage.  Ideally, the ratio of lactic acid to acetic acid should not be less than 3:1 and higher is better.  Poor ratios of lactic acid to acetic acid indicate that you should consider using a microbial inoculant containing homolactic acid bacteria when ensiling.  Determination of individual acid content is usually accomplished by some type of chromatography.  However, some labs use enzymatic methods to detect lactic acid.  In this case, the L-form of lactate is determined and then multiplied by 2 to obtain an estimate of total lactate (since the L- and D- forms are usually present in a 1:1 ratio).

 

Ethanol can also be found in some silage.  In corn silage, ethanol is between 1 to 3% of the DM but can sometimes be greater than 6 - 7%.  The ethanol content in grass and legume silages is usually less than 1.5%.  Silages that have high levels of ethanol have usually undergone an extensive fermentation due to yeasts.  Although the energy value of such silages is generally good, alcoholic fermentations result in large losses of nutrients.  Such silages will also have a tendency to heat rapidly when exposed to air and bunk life will be short.  Silages preserved with acids (as in Europe) often have high ethanol levels, have high yeast counts, and are very unstable when exposed to air.  Ethanol is usually determined by a chromatographic or enzymatic method in the lab.

 

Ammonia concentration is usually expressed as a percentage of the crude protein (CP) or nitrogen content.  In corn silage, ammonia is normally between 5 to 7 % of the CP but it is usually higher (10 to 15%) in grass and alfalfa silage.  High concentrations of ammonia are an indicator of extensive protein degradation.  These silages often have high contents of soluble protein.  Protein degradation occurs extensively in wet silages (< 30 to 35% DM) and can be due to plant degrading enzymes and/or undesirable clostridial microorganisms.  A longer wilt (higher DM%) may be called for to prevent this occurrence.  Ideally, ammonia should be <7% of the protein content but this is difficult to achieve in high protein forages (e.g. alfalfa).  Ammonia can be determined by colorimetric or distillation procedures or with an ammonia-specific ion probe.

 

In addition to determining chemical products, some analytical labs will enumerate the numbers of yeasts and molds in silages.  Yeast and molds are determined by growing them on a selective medium.  The number of colonies formed on a petri plate with this media are counted and expressed as colony forming units (cfu) per gram of wet silage.  High numbers of yeast and molds are usually an indication of poor packing of a feed or a too slow feed out rate that exposes silage to oxygen.  Silages with more than 100,000 (or 1 ´ 105 cfu per gram of silage are usually prone to rapid heating and poor bunk life because many yeasts degrade lactic acid and are the cause for spoiling when silages are exposed to air.

Some precautions should be taken when sending silage samples to a laboratory for fermentation analyses as samples that are delayed in transit and subjected to warm temperatures may start to degrade before they reach the lab.  Never send a sample on a Friday and opt for a next day delivery if possible.  Silage samples may be refrigerated or frozen before shipping to aid in preservation during transit.  Samples for the determination of yeasts and molds should only be refrigerated, not frozen.  Include ice packs and place in a shipping cooler if practical.  In addition, if you can, dig deep (10 to 12 inches) into the silage mass to retrieve a sample for analyses.  Avoid taking samples that have been exposed to oxygen as they may have deteriorated and not be representative of what is being fed.  Table 1 shows some typical fermentation products for the more commonly used silages.

 

Table 1.  Amounts of common fermentation end products in various silages.

 

Item	Alfalfa Silage, 30 - 35% DM	Alfalfa Silage, 45 - 55% DM	Grass Silage, 25 - 35%  DM	Corn Silage, 35 - 40% DM	HM Corn, 75% DM
pH	4.3 - 4.5	4.7 - 5.0	4.3 - 4.7	3.7 - 4.2	4.0 - 4.5
Lactic acid, %	7 - 8	2 - 4	6 - 10	4 - 7	0.5 - 2.0
Acetic acid, %	2 - 3	0.5 - 2.0	1 - 3	1 - 3	< 0.5
Propionic acid, %	< 0.5	< 0.1	< 0.1	< 0.1	< 0.1
Butyric acid, %	< 0.5	0	0.5 - 1.0	0	0
Ethanol, %	0.5 - 1.0	0.5	0.5 - 1.0	1 - 3	0.2 - 2.0
Ammonia-N, % of CP	10 - 15	< 12	8 - 12	5 - 7	< 10

 

 

Management factors markedly influence silage fermentation quality.  Good fermentations require that the forage be harvested at an immature stage of growth, that it be chopped to the correct particle size to aid packing, and that the silo be filled quickly to reduce plant respiration of the sugars.  The silo must also be firmly packed to remove oxygen, and the forage must be properly covered with plastic.  In bunk and pit silos, the plastic cover must be completely weighted down with tires to minimize air infiltration and spoilage.

 

Of the fermentation end products that are normally measured in research studies, only silage pH can be easily measured on the farm.  More often than not, producers can smell problem silages.  For example, high levels of ethanol produced by yeasts can be readily detected.  Similarly, butyric acid silage smells rank, rancid, and sometimes fishy (due to protein degradation).  Routine analyses for silage fermentation end-products is costly, probably unnecessary and unfortunately, analyses for most fermentation end-products cannot be used directly to assist in balancing a diet for a cow.  However, select samples should be submitted for analyses, as they may be an indication of the type of fermentation that occurred.