Investigation of liquid-solid hydrodynamic boundary layers and oxygen requirements in hairy root cultures

Abstract

Rates of oxygen uptake, growth and alkaloid production by hairy roots in submerged culture were investigated using a recirculation reactor allowing operation at high liquid velocities for removal of hydrodynamic boundary layers. Measurements were performed at dissolved oxygen tensions of 31–450% air saturation. Critical oxygen concentrations for Atropa belladonna hairy roots were above air saturation, viz. 100–125% air saturation for oxygen uptake and 150% air saturation for growth, demonstrating that these roots cultivated in reactors with air sparging are oxygen‐limited. The critical oxygen tension for oxygen uptake by Solanum aviculare hairy roots was 75% air saturation. Both the specific oxygen uptake rate and specific growth rate of A. belladonna hairy roots were dependent on the mass (g dry weight) of roots present; even in the absence of boundary layers, growth did not remain exponential over the entire culture period. Cryo‐scanning electron microscopy showed that hairy roots grown submerged in liquid medium were covered with thick layers of hydrated mucilage and root hairs, representing a significant additional barrier to oxygen transfer. Roots protruding out of the liquid medium showed no evidence of mucilage accumulation. The specific oxygen demand of A. belladonna root tips was 3.3–11.5 times higher than for the remainder of the roots, the ratio increasing as the dissolved oxygen tension was reduced. Specific growth rates, biomass yields from sugar, and atropine levels were maximum at around 150% air saturation, but decreased significantly with oxygen concentrations above ca. 200%. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 64: 729–740, 1999.

Modelling in vitro growth of dense root networks

“…Hairy root cultures, producing many of the same important secondary metabolites as the whole plant, are a potential means for producing valuable plant compounds (Williams and Doran, 1999). Hairy roots are obtained through transformation by Agrobacterium rhizogenes and are special in the sense that these plants lack shoots and are composed mainly of a dense growing root system (Fig.…”

Strategies for Enhancing Monoclonal Antibody Accumulation in Plant Cell and Organ Cultures

“…Similar enhancement of antibody heavy-chain accumulation has been reported in tobacco cell suspensions with improved oxygen transfer (42). The observed beneficial effects of operating hairy root cultures at oxygen tensions above ambient are consistent with the relatively high critical oxygen tensions measured for hairy roots (15,16), reflecting the unavoidable development of significant oxygen concentration gradients within the root tissues.…”

The growth of single roots ofArtemisia annua in nutrient mist reactors

“…Shear also decreases root elongation (Hitaka et al, 1997) and the viability of root tips (Kino-Oka et al, 1999). In addition, Williams and Doran (1999) showed that roots grown in liquid produce a much thicker layer of mucilage than roots grown in moist air.…”

“…A proliferation of root tips compared with elongated cells can increase oxygen demand tenfold (Ramakrishnan and Curtis, 1995;Williams and Doran, 1999) and the extra aeration required to meet the oxygen demand of the growing roots in a liquid-phase reactor becomes more difficult because the pressure drop across the reactor also increases with increased branching. The problem is compounded if the extra oxygen stimulates root hair formation and further increases fluid drag.…”

“…The problem is compounded if the extra oxygen stimulates root hair formation and further increases fluid drag. Finally, the mucilage present on roots grown in liquid can supply additional resistance to O 2 transfer (Williams and Doran, 1999). From the standpoint of metabolite production, root tips may produce more (Wobbe et al, 1997) secondary products than the mature root, and lateral roots may produce more products than primary roots (Woo et al, 1997).…”