Beer has been brewed for thousands of years and for the majority of this time has been produced using mixed culture fermentation via a complex spectrum of microflora, not just brewing yeast. Indeed, arguably, sourness (to some extent) has been an important and prevalent characteristic of beer throughout history. It is only in the last 500-600 years that hops have been used as the widespread and dominating flavouring of beer, with their bacteriostatic properties inhibiting the presence and activity of some bacteria in particular. In more recent history, the pioneering work of scientists such as Pasteur and Hansen saw the development of sterile culturing techniques and the isolation and purification of single cell cultures. This, coupled with technological developments in the mid-late Victorian period, saw a move away from mixed cultures and the rise of pure culture fermentations and greater homogeneity in beer which still dominates the global landscape (bearing in mind that lager/pilsner production accounts for 90%+ of world beer production).
However, pockets of unique and iconic beers using mixed microflora remained in Europe. Sour beer is certainly nothing new. We would perhaps associate sour beers chiefly with Belgium with styles such as Lambic, Flanders Red, Gose etc but so to could sour beers be found in Germany (Berliner Weisse) and the UK (Oak aged ales). Fast forward to modern day and we are now seeing an explosion in the popularity of sours, nothing short of a renaissance. This, like most phenomenon in the modern craft beer movement, is being fueled and influenced by the US. In the 1990s beer imports saw an influx of Belgian beers into the US which had a profound effect on brewers and consumers alike. 2002 saw the first time sours were entered as a standalone category at the great American Beer Festival (with only a handful of entries) but since then great expansion and popularity in sour beers now sees hundreds of entries. The diversity in sour styles, flavours and creativity is now going global and having been influenced by iconic European styles and techniques modern brewers worldwide are taking sours to exciting new places which is being reflected in demand for these beers.
Lactobacillus can largely be considered the primary souring bacteria and has a diverse range of subspecies (see R&D trials later in the article). Lactobacillus is at the heart and the dominating characteristic of sour beers such as Berliner Weisse and Gose but is also used as part of mixed fermentation in many sour styles. Lactobacillus produces lactic acid very rapidly, imparting a soft and tangy flavor. Temperature sensitivity is crucial for performance and this does vary by subspecies, 30-49°C being a common temperature range for Lactobacillus activity. They are typically very sensitive to hops (though again this is species dependent) and as little as c.8 IBU can inhibit growth and activity. Lactobacillus can be categorised as heterofermentative (producing lactic acid and other byproducts such as CO2 and Ethanol) and homofermentative strains which produce lactic acid alone.
Pediocococcus is also a common souring bacteria but by contrast to lactobacillus is much slower (potentially taking months to reach lower pH levels) which may influence the technique selected to sour. Although slower in activity it is more resistant to hops as well as acids and thus can achieve pH levels of 3.0 and lower (Lactobacillus typically achieving 3.2-3.5 pH). The result being that Pedicoccus produces a much harsher and sharper taste as compared to Lactobacillus. Most species will produce diacetyl to varying concentrations, which is largely considered a negative by many brewers and in the right conditions can produce exopolysaccharides resulting in “sick” or “ropey” beer.
Brettanomyces, wild yeast and not bacteria, is often used to ferment sour beers. Unlike common brewing yeasts (S. cerevisiae and S. pastorianus) Brettanomyces can utilize a broad range of sugars including dextrin material but is typically slower. A common misconception of Brettanomyces is that it contributes to acidity similar to bacteria. It does not on its own but is often used alongside bacteria. Depending on the subspecies Brettanomyces can produce a diverse range of esters, phenols and other compounds resulting in flavours that lend themselves well to sour beer styles. For example, B.Bruxellenis tends to produce more earthy, woody and musty notes versus fruity, pineapple esters that would be associated with B.Claussenii.
Sources of Lactic Acid producing bacteria
In searching for preferable bacteria used for sour beer production brewers turn to many sources to achieve desired results. Some of the most common include:
Laboratory – Commercially available strains via laboratories are becoming increasingly available either as a pure or mixed culture (more common).
Bottle Cultures – Brewers and microbiologists harvest cultures found in the sediment/dregs of unpasteurized sour beers and then grow these cultures up. These would typically be mixed and often complex cultures.
Nature – Exposing wort or beer to atmosphere and allowing naturally present bacteria and wild yeasts to sour. A traditional technique especially favored in Belgium.
Yoghurt – A range of dairy products including yoghurt are fermented with Lactobacillus and adding yoghurt containing a spectrum to wort of beer has been used in sour beer production.
Un-mashed grains – Lactobacillus is often present on the grains/cereals used in brewing and the addition of crushed and un-mashed grains in the brew house can be used as a technique for souring.
Techniques for souring
• Liquor, grain adjustment
• Bacteria from grain or inoculated
• 2 – 3 days
• Wort inoculated with LAB
• 2 -3 days
• Mixed sacc, LAB & Brett
• Typical fermentation time
• Often in wood (or Keolschip)
• Mixed spectrum of microflora
• Greater complexity
Typical/example kettle sour process
With the increased consumption of sour beers (containing lactic acid) comes a demand to be able to produce such beers in a convenient and controlled way such as using dried bacteria in pitchable sizes. Based on encouraging results with a L. plantarum strain it was decided to evaluate a wider range of available dried lactic acid bacteria. The target is to achieve pH 3.5 or lower within 48 hours of fermentation with high lactic acid and low acetic acid concentrations. Here we test several lactic acid bacteria strains. L. plantarum was included as a control because it was the best performer in previous trials.
Sour beers are becoming more popular in the market today and brewers looking for an easy way to produce this beer style without propagating and maintaining their own lactic acid bacteria cultures. Using dried bacteria cultures in pitchable sizes would be a convenient solution. 6 lactic acid bacteria strains were fermented in 12 % unhoped malt extract at four different temperatures. L helveticus and L. acidophilus showed the highest activity at 40 ºC, which resulted in the highest lactic acid concentration. The highest acetic acid concentrations were produced at 20 ºC and in general decreased with increasing fermentation temperatures. L helveticus and L. acidophilus seem suitable candidates for sour beer production. L. delbrueckii might be an interesting addition to the portfolio of lactic acid bacteria for sour beer production because it produced some interesting fruity notes.
Materials and Methods
Beer fermentations with the samples were performed at 4 different temperatures (20 ºC, 30 ºC, 40 ºC and 50 ºC in 500ml media bottles. The wort was prepared from malt extract to 12° Plato and transferred into sterile bottles. Bacteria were rehydrated at room temperature for 15 minutes and pitched at 1g/hl except for L. plantarum (strain A) which was pitched at 10 g/hl as recommended. Daily measurements of gravity and pH were taken over the course of the fermentation.
Samples were taken at the end of each fermentation and analyzed for lactic acid, acetic acid and glycerol. The analysis was performed by HPLC with a column Jolie Waters Ic-Pak Ion-exclusion 50A 7um 7.8X150mm
Both Lactobacillus plantarum strains showed the highest activity at 20 ºC and 30 ºC resulting in the fastest pH drop and lowest pH after 3 days fermentation. At 30 ºC and 40 ºC all strains reached the target pH of 3.5 within 2 days with the exemption of L. brevis strain (30 ºC & 40 ºC) and L. delbrueckii (30 ºC). L. helveticus and L. acidophilus showed the highest activity at 40 ºC and were still active at 50 ºC whereas all other strains were almost inactive at that temperature (graphs 1 – 4).
HPLC results indicate that the highest lactic acid concentrations were produced at 40 ºC by L. helveticus followed by L. acidophilus. At 30 ºC all strains produced similar high concentrations of lactic acid. L brevis is the most sensitive strain to higher fermentation temperatures producing the highest concentration at 20 ºC (graph 5). The highest acetic acid concentrations were produced at 20 ºC and in general decreased with increasing fermentation temperatures (graph 6). The highest glycerol concentrations were measured at 30 ºC produced by L. plantarum and L helveticus (graph 7).
The fermentations were tasted after 3 days by a tasting panel. In general the fermentations with L helveticus and L. acidophilus were described as having the most intense sour taste and smell. L. delbrueckii produced some interesting fruity notes. One of the two bottles of L helveticus at 30 ºC produced a biofilm and smelled like “sweaty socks” and had a roasted aftertaste.
Sensory summary of fermentations
The production of sour beers is fast becoming increasingly prevalent and the requirement for reliable and consistent techniques and desirable flavor profile is highly relevant. The diversity in Lactobacillus sub species is evident in terms of performance, temperature sensitivity and optimal conditions. By identifying, characterizing and understanding how these subspecies work and moreover what techniques for souring they are best suited for continuing research and development of easy to handle high performance bacteria cultures can be of benefit to brewers producing sour beers. A number of strains available in the Lallemand culture collection and produced in freeze dried form appear to be ideal candidates for application in brewing.