Is enzymatic oligonucleotide production ready for commercial-scale manufacturing?

Today, most oligonucleotides are made using solid-phase phosphoramidite chemistry, a method that has been the industry standard since the 1980s. It is reliable, automated, and well-characterised. But it also has limitations, including high solvent consumption, significant material costs, increasing process mass intensity (PMI) concerns, and difficulties scaling beyond modest batch sizes. As a result, innovators across academia and industry are investigating enzymatic synthesis as a greener, potentially more scalable alternative.

Why enzymatic oligonucleotide synthesis is gaining commercial interest

Unlike traditional solid-phase synthesis, enzymatic approaches build oligos biocatalytically, using enzymes to add nucleotides in aqueous conditions. Several advantages immediately appear: 

• Lower solvent usage. 

• Fewer hazardous reagents. 

• Potentially reduced purification burden. 

• Gentler reaction conditions. 

• More flexibility on sequence length.

This isn’t a trivial manufacturing decision anymore. The global oligonucleotide synthesis market is projected to grow from around USD 10.5 billion in 2025 to USD 24.7 billion by 2030, a compound annual growth rate close to 20%. That means any inefficiency in upstream or downstream processing will soon be magnified at industrial volume. 

For many companies, the sustainability and safety benefits alone justify exploration. But the real prize is scalability. 

At CPI’s Oligonucleotides Symposium in 2025, Professor Glenn Burley from the University of Strathclyde highlighted why enzymatic methods are attracting serious interest. Describing nucleoside transglycosylase enzymes (NDTs), he noted that: 

“Enzymatic synthesis is a really nice method… these enzymes are robust, tolerant to solvents, and allow gram scale synthesis in batch or flow.”

The keyword there is flow. Flow-compatible enzymatic synthesis opens the door to genuine continuous manufacturing of oligonucleotides, a feat that solid-phase chemistry has never achieved. 

Why sustainability and PMI are driving enzymatic approaches

A recent comparative assessment estimated a PMI of approximately 530 kg of raw material per kg of oligonucleotide for an enzymatic route, compared with well over 4,000 kg/​kg for traditional phosphoramidite synthesis. That is almost an order-of-magnitude improvement in material use for the same output. 

And across multiple development programs, PMI for synthetic oligo production is typically reported in the 3,000 – 7,000 range, far higher than most small molecules (often <50).

Environmental factors, regulatory pressure, and supply-chain fragility all make enzymatic approaches compelling. 

The bottlenecks and unanswered questions

Despite the enthusiasm for this this innovative technology, significant challenges remain. Enzymatic oligo production is still largely pre-commercial. Most success stories involve bench-scale research, not GMP processes producing kilograms of active pharmaceutical ingredient.

Unlike small molecules or biologics, there is no clear roadmap yet for enzyme selection, immobilisation, purification, recycling, or stepwise activation for long or complex oligos. And while enzymatic methods may reduce environmental impact, they introduce their own complexities, including limited enzyme availability, sequence-dependent performance, uncertainty in the patent landscape, and downstream analytics that differ from established synthetic approaches. 

Hybrid models may get to scale first

One approach gaining traction is the hybrid model, in which enzymes produce short, high-fidelity fragments that are later chemically ligated. This could significantly reduce the number of synthetic steps, decrease solvent usage, and simplify impurity profiles.

The quiet enabling factor: manufacturing testbeds

The step from research-scale enzymatic methods to commercial oligo production will not be determined in academic papers alone. The real breakthrough will come through process benchmarking, comparative studies, and GMP-relevant validation. 

This is where neutral, innovation-led manufacturing environments are important. Organisations such as CPI offer facilities where emerging enzymatic approaches can be evaluated alongside legacy solid-phase methods, run under manufacturing-like conditions, linked to downstream purification development, and supported with analytics and process control technologies. 

Despite rapid technological advances, the oligonucleotide market still has more questions than answers. Sponsors, developers and investors are grappling with real uncertainty around how emerging enzymatic and hybrid synthesis routes will scale, how impurity profiles will shift with new unit operations, and what the true total cost of goods and sustainability implications are when compared with established phosphoramidite methods. 

CPI is uniquely positioned to turn these questions into data: benchmarking alternative routes, carrying out rigorous techno-economic analysis, and developing scalable downstream and validated analytical strategies that make disparate data points directly comparable. Instead of speculating about ​“what might work,” project teams can partner with CPI to generate datasets that clarify risk and inform tactical decisions, whether for early-stage process feasibility, GMP-aligned scale-up, or regulatory filings. 

A realistic outlook

If we look at long-term industry needs such as reduced PMI, lower solvent load, and greener chemistry, then enzymatic synthesis ticks almost every box. It’s not yet a replacement for phosphoramidite chemistry, but it is certainly a future pillar in oligo manufacturing capacity. 

The companies that start learning now will be the ones ready when enzymatic systems reach maturity. And those who combine experimentation with manufacturing partners will have the best chance of getting there first. 

Turn uncertainty into data-driven decisions

If you’re exploring enzymatic or hybrid oligonucleotide manufacturing or need independent validation to support scale-up, investment, or regulatory decisions, contact CPI to discuss how our manufacturing testbeds, analytics, and techno-economic expertise can support you.

Let’s innovate together.